Akira Yamazaki; Ao Takezawa; Kazusa Nishimura; Ko Motoki; Kyoka Nagasaka; Ryohei Nakano; Tetsuya Nakazaki; Munetaka Hosokawa
The Horticulture JournalJapanese Society for Horticultural Science2189-01022024
Kenta Shirasawa; Tomoya Esumi; Akihiro Itai; Katsunori Hatakeyama; Tadashi Takashina; Takuji Yakuwa; Katsuhiko Sumitomo; Takeshi Kurokura; Eigo f*ckai; Keiichi Sato; Takehiko Shimada; Katsuhiro Shiratake; Munetaka Hosokawa; Yuki Monden; Makoto Kusaba; Hidetoshi Ikegami; Sachiko Isobe
Cold Spring Harbor Laboratory2023/07
AbstractFlowering cherry cultivar ‘Somei-Yoshino’ (Cerasus×yedoensis) has been clonally propagated and spread all around the world including Japan. ‘Somei-Yoshino’ is thought to be an interspecific hybrid derived fromC. spachianaandC. speciosa; however, its origin is unclear. Since somatic mutations are randomly induced in genomes and stably transmitted through generations, we aimed to identify somatic mutations in the genome of ‘Somei-Yoshino’ to trace its propagation path. A total of 46 ‘Somei-Yoshino’ clones were collected from all over Japan and subjected to whole-genome sequencing. The results revealed 684 single nucleotide variants, of which 71 were found in more than two clones. Clustering analysis of the 46 clones using these 71 variants revealed six groups, four of which contained 40 of the 46 clones. In addition, because each of the four clones closely planted in Ueno Park, Tokyo, Japan, clustered into the four different groups, we considered that these four clones could be the ancestors of the ‘Somei-Yoshino’ clones found in Japan. Furthermore, based on the comparison of mutant alleles with the genomes ofCerasusspecies, one of the four trees was concluded as the closest to the origin. Here, we propose that the origin of ‘Somei-Yoshino’ is a chimera derived from at least four somatic mutants.
Yu Kinosh*ta; Ko Motoki; Munetaka Hosokawa
Theoretical and Applied GeneticsSpringer Science and Business Media LLC136(3)0040-57522023/03
Akira Yamazaki; Ao Takezawa; Ryohei Nakano; Kazusa Nishimura; Ko Motoki; Munetaka Hosokawa; Tetsuya Nakazaki
Journal of Horticultural ResearchWalter de Gruyter GmbH30(2)105 - 1162023/01
Abstract “Autonomous fruit set” refers to self-pollination and fruit set without pollen vectors such as vibration or insects. Autonomous fruit set under high-temperature stress is an important breeding goal as climate change can reduce fruit yields in Capsicum. We screened Capsicum cultivars for autonomous fruit set ability in a greenhouse environment and investigated pollen germination, viability, pollen grains number, chlorophyll fluorescence (Fv/Fm), style length, anther cone length, and anthesis stage under high temperatures in order to identify indicator traits for screening more genotypes with autonomous fruit set ability. The fruit set of the ‘Takanotsume’ (57.7 ± 20.6%) and ‘Goshiki Kyokko’ (52.2 ± 14.2%) cultivars (both C. annuum) were higher than those of other cultivars. Correlation analysis showed that pollen germination had the highest correlation with fruit set in C. annuum cultivars (r = 0.63). These results indicate that ‘Takanotsume’ and ‘Goshiki Kyokko’ are useful cultivars for novel breeding programs focusing on autonomous fruit sets under high temperatures, and pollen germination in C. annuum was a convincing candidate for an indicator trait of autonomous fruit set ability under high temperatures.
Ayumu Kono; Ayumu Kawabata; Akira Yamazaki; Yuma Ohkubo; Adriano Sofo; Munetaka Hosokawa
The Horticulture JournalJapanese Society for Horticultural Science2189-01022023
Ko Motoki; Yu Kinosh*ta; Ryohei Nakano; Munetaka Hosokawa; Tetsuya Nakazaki
The Horticulture JournalJapanese Society for Horticultural Science92(1)66 - 762189-01022023
Kenta Shirasawa; Munetaka Hosokawa; Yasuo Yasui; Atsushi Toyoda; Sachiko Isobe
Cold Spring Harbor Laboratory2022/09
AbstractHere, we report the genome sequence of a popular Japanese chili pepper landrace, Capsicum annuum ‘Takanotsume’. We used long-read sequencing and optical mapping, together with the genetic mapping technique, to obtain the chromosome-scale genome assembly of ‘Takanotsume’. The assembly consists of 12 pseudomolecules, which corresponds to the basic chromosome number of C. annuum, and is 3,058.5 Mb in size, spanning 97.0% of the estimated genome size. A total of 34,324 high-confidence genes were predicted in the genome, and 83.4% of the genome assembly was occupied by repetitive sequences. Comparative genomics of linked-read sequencing-derived de novo genome assemblies of two Capsicum chinense lines and whole-genome resequencing-derived genome assemblies of Capsicum species revealed not only nucleotide sequence variations but also genome structure variations (i.e., chromosomal rearrangements) between ‘Takanotsume’ and its relatives. Overall, the genome sequence data generated in this study will accelerate the pan-genomics and breeding of Capsicum, and facilitate the dissection of genetic mechanisms underlying the agronomically important traits of ‘Takanotsume’.
Sho Ohno; Haruka Yamada; Kei Maruyama; Ayumi Deguchi; Yasunari Kato; Mizuki Yokota; Fumi Tatsuzawa; Munetaka Hosokawa; Motoaki Doi
PlantaSpringer Science and Business Media LLC256(3)0032-09352022/09
Ko Motoki; Yu Kinosh*ta; Ryohei Nakano; Munetaka Hosokawa; Tetsuya Nakazaki
Plant and Cell PhysiologyOxford University Press (OUP)63(9)1230 - 12410032-07812022/07
Abstract Grafting-induced flowering is a key phenomenon to understand systemic floral induction caused by florigen. It can also be used as breeding technique enabling rapid seed production of crops with long generation times. However, the degree of floral induction in grafted plants is often variable. Moreover, it is difficult in some crop species. Here, we explored the factors promoting variability in the grafting-induced flowering of cabbage (Brassica oleracea L. var. capitata), an important vegetable crop with a long generation time, via the quantitative analysis of florigen accumulation. Significant variability in the flowering response of grafted cabbage was observed when rootstocks of different genotypes were used. As reported previously, B. oleracea rootstocks didn’t induce flowering of grafted cabbage plants, but radish (Raphanus sativus L.) rootstocks unstably did, depending on the accessions used. Immunoblotting analysis of the FLOWERING LOCUS T (FT) protein, a main component of florigen, revealed that floral induction was quantitatively correlated with the level of accumulated FT protein in the grafted scion. To identify rootstock factors that cause variability in the floral induction of the grafted scion, we investigated FT protein accumulation and flowering response in grafted scions when the transcription levels of FT and the leaf area of rootstocks were altered by vernalization, daylength, and leaf trimming treatments. We concluded that increasing the total amount of FT protein produced in the rootstock is important for stable floral induction of the grafted cabbage, and this can be accomplished by increasing FT transcription and the leaf area of the rootstock.
Emi Ota; Fumihiro Nishimura; Mitsutaka Mori; Masaya Tanaka; Takeshi Kanto; Munetaka Hosokawa; Masahiro Osakabe; Mamoru Satou; Minoru Takesh*ta
Plant PathologyWiley70(6)1378 - 13870032-08622021/04[Refereed]
Tamanna Abedin; Atsushi Yamamoto; Takahiro Hayashi; Munetaka Hosokawa
Scientia HorticulturaeElsevier {BV}276109604 - 1096040304-42382021/01
Yu Kinosh*ta; Ko Motoki; Munetaka Hosokawa
The Horticulture JournalJapanese Society for Horticultural Science90(4)374 - 3812189-01022021
Akira Yamazaki; Kenta Shirasawa; Munetaka Hosokawa
EUPHYTICASPRINGER216(11)0014-23362020/10
Breeding for thermotolerance in processes determining fruit set and pollen germination is increasingly needed as global warming progresses. We previously reported good yields at high temperatures from the F-1 hybrid of the Capsicum chinense cultivars 'Sy-2' and 'No. 3686', both of which yield poorly at high temperatures. Moreover, the percentages of fruit set at high temperature in the F-2 population segregated quantitively. In addition, percentages of fruit set in some F-2 individuals exceeded those of the F-1 hybrid. Analysis of the F-2 population using double-digest restriction site-associated DNA sequencing (ddRAD-Seq) shows that the percentage of fruit set under high temperature is strongly associated to single nucleotide polymorphisms (SNPs) in a certain gene region of chromosome 6 and weakly associated to a gene region on the chromosome 3. Furthermore, the pollen germination rate is strongly associated to SNPs of the same regions on chromosomes 3 and 6. Analysis of the genotypes and traits in another F-2 population shows that the genotype determined by the cleaved amplified polymorphic sequence marker designed on the candidate gene region is associated to the percentage of fruit set and the pollen germination rate at high temperatures, supporting the results of ddRAD-Seq. The presence of alleles from both the gene regions of chromosome 3 from 'No. 3686' and chromosome 6 from 'Sy-2' increased the percentage of fruit set additively. We suggest that genes controlling fruit set and pollen germination at high temperature are located in these two gene regions.
Daichi Tomimori; Munetaka Hosokawa; Shinichi Aoki; Masahiro Osakabe
Environmental entomologyOxford University Press (OUP)49(4)886 - 8940046-225X2020/08[Refereed]
Perilla, Perilla frutescens (L.) Britton var. crispa (Thunb.) H. Deane, is traditionally cultivated as an edible/medicinal crop in East Asia. Its essential oil contains many bioactive compounds that are expected to have high pharmacological functionality, as well as antimicrobial and insecticidal activity. Spider mites are a major pest group for perilla cultivation. The two-spotted spider mite, Tetranychus urticae Koch, possesses divergent detoxification enzymes and has developed resistance against most acaricides. The essential oil content of perilla halves from the pre-flowering phase to the flowering phase, and ultraviolet (UV)-B radiation generally increases defense compounds. To clarify the effects of this change in essential oil content and the effects of UV-B pretreatment, we investigated the developmental success and egg production of T. urticae on leaves from the preflowering and flowering phases cultivated with and without nighttime UV-B irradiation. Both the parameters significantly increased on leaves from the flowering phase in comparison with that from the preflowering phase, suggesting that constitutively produced essential oil provided protection against mite pests in a growth phase-specific manner. The defense system also extended the developmental period of mites on red perilla leaves, but not on green perilla leaves, in preflowering phase. Although egg production was lower on red perilla leaves pretreated with UV-B, no negative effects were caused on the developmental success and duration on red and green perilla and the egg production on green perilla by UV-B pretreatment. Our findings reveal a significant impact of investment allocation of perilla plants and a small contribution of UV-B irradiation to the plant defense system.
Hiroshi Okada; Tamanna Abedin; Atsushi Yamamoto; Takahiro Hayashi; Munetaka Hosokawa
Scientia HorticulturaeElsevier {BV}267109252 - 1092520304-42382020/06
Increased percentage of fruit set of F1 hybrid of Capsicum chinense during high-temperature period.
HOSOKAWA Munetaka
Sci. Hortic.243421 - 4272019[Refereed]
HOSOKAWA Munetaka
Sci. HorticElsevier BV2441 - 90304-42382019[Refereed]
Tomoyuki Nabeshima; Yosuke Matsush*ta; Munetaka Hosokawa
Viruses10(12)729 - 7292018/12[Refereed]
Chrysanthemum stunt viroid (CSVd) is one of the most severe threats in Chrysanthemum morifolium production. Over the last decade, several studies have reported the natural occurrence of CSVd resistance in chrysanthemum germplasms. Such CSVd-resistant germplasms are desirable for the stable production of chrysanthemum plants. Current surveys include finding new resistant chrysanthemum cultivars, breeding, and revealing resistant mechanisms. We review the progress, from discovery to current status, of CSVd-resistance studies, while introducing information on the improvement of associated inoculation and diagnostic techniques.
Sho Ohno; Wakako Hori; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
PlantaSpringer Verlag247(2)413 - 4281432-20482018/02[Refereed]
MAIN CONCLUSION: Post-transcriptional gene silencing (PTGS) of a chalcone synthase ( DvCHS2 ) occurred in the white part of bicolor petals and flavonoid-poor leaves; however, it did not in red petals and flavonoid-rich leaves. Petal color lability is a prominent feature of bicolor dahlia cultivars, and causes plants to produce not only original bicolor petals with colored bases and pure white tips, but also frequently single-colored petals without white tips. In this study, we analysed the molecular mechanisms that are associated with petal color lability using the red-white bicolor cultivar 'Yuino'. Red single-colored petals lose their white tips as a result of recover of flavonoid biosynthesis. Among flavonoid biosynthetic genes including four chalcone synthase (CHS)-like genes (DvCHS1, DvCHS2, DvCHS3, and DvCHS4), DvCHS1 and DvCHS2 had significantly lower expression levels in the white part of bicolor petals than in red petals, while DvCHS3, DvCHS4, and other flavonoid biosynthetic genes had almost the same expression levels. Small RNAs from the white part of a bicolor petal were mapped onto DvCHS1 and DvCHS2, while small RNAs from a red single-colored petal were not mapped onto any of the four CHS genes. A relationship between petal color and leaf flavonoid accumulation has previously been demonstrated, whereby red petal-producing plants accumulate flavonoids in their leaves, while bicolor petal-producing plants tend not to. The expression level of DvCHS2 was down-regulated in flavonoid-poor leaves and small RNAs from flavonoid-poor leaves were mapped onto DvCHS2, suggesting that the down-regulation of DvCHS2 in flavonoid-poor leaves occurs post-transcriptionally. Genomic analysis also suggested that DvCHS2 is the key gene involved in bicolor formation. Together, these results suggest that post-transcriptional gene silencing of DvCHS2 plays a key role in phenotypic lability in this bicolor dahlia.
Sho Ohno; Wakako Hori; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
Horticulture JournalJapanese Society for Horticultural Science87(1)140 - 1482189-01102018[Refereed]
Bicolor flowering dahlias are a group of cultivars that produce inflorescences with bicolored petals characterized by a colored basal part and a white tip. However, they frequently produce single-colored petals, even if they are vegetatively propagated. In a previous study, strong relationships between inflorescence color and leaf phenotype were observed in a red–white bicolor flowering dahlia ‘Yuino’ red petal-producing individuals accumulate flavonoids in leaves, whereas only bicolor petal-producing individuals tend not to accumulate them in leaves. Flavonoids in leaves are assumed to be chalcones. In this study, we investigated flavonoids in the leaves of ‘Yuino’ by nuclear magnetic resonance analysis and identified six caffeoyl esters, four flavonol derivatives, and three novel butein derivatives in the flavonoid-rich leaves of ‘Yuino’. The three novel compounds were butein 4',4-O-di-[2-O-(β-glucopyranosyl)-β-glucopyranoside], butein 4'-O-[2-O-(β-glucopyranosyl)-β-glucopyranoside]-4-O-β-glucopyranoside, and butein 4'-[6-O-(3-hydroxy-3-methylglutaryl)-β-glucopyranoside]-4-O-β-glucopyranoside. On the other hand, only caffeoyl esters were detected in flavonoid-poor leaves. These data demonstrated that flavonoid-rich leaves accumulated the flavonoids of butein and flavonol derivatives. The common enzyme for the biosynthesis of butein and flavonol derivatives is chalcone synthase thus, the importance of chalcone synthase for phenotypic lability in ‘Yuino’ was confirmed.
Ko Motoki; Yu Kinosh*ta; Munetaka Hosokawa
Frontiers in plant science91967 - 19672018[Refereed]
Cabbage (Brassica oleracea var. capitata) requires a long-term low-temperature exposure for floral induction, causing a delay in the breeding cycle. The objective of this study is to develop a method to induce flowering in cabbage without low-temperature treatment, using a grafting method. We conducted grafting experiments using two flower-induced Chinese kale cultivars (B. oleracea var. alboglabra) and seven radish cultivars/accessions as rootstocks and investigated the flowering response of grafted cabbage scions without low-temperature treatment. "Watanabe-seiko No.1" cabbage, when grafted onto the two Chinese kale cultivars, did not formed flower buds. Flowering was successfully induced in "Watanabe-seiko No.1" by grafting onto three out of the seven tested radish cultivars, and in "Kinkei No.201" and "Red cabbage" by grafting onto one tested radish cultivar. In "Watanabe-seiko No.1," the earliest flower bud appearance was observed at 29 days after grafting. Seeds were also obtained from the three cabbage cultivars that flowered by grafting. Gene expression analysis of "Watanabe-seiko No.1" cabbage scions which formed flower buds by grafting, revealed high expression of the hom*olog of the floral integrator, SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (BoSOC1), at the time of flower bud appearance. However, in the same leaf samples, we observed low expression of two hom*ologs of florigen, FLOWERING LOCUS T (BoFT.C2 and BoFT.C6). In addition, two hom*ologs of the floral repressor FLOWERING LOCUS C (BoFLC3 and BoFLC4), which are known to be down-regulated before flower bud differentiation in the vernalization pathway, were highly expressed, indicating that grafting onto radish induces cabbage flowering independently of the vernalization pathway. The expression level of the radish FT hom*olog (RsFT) in "Rat's tail-G2," which had highly induced flowering in the grafted cabbage scion, was higher than in the other radish cultivars. However, although "Rat's tail-CH" effectively induced flowering in the cabbage scion, the expression of RsFT was low in this cultivar. In this study, floral induction of non-vernalized cabbage cannot be explained by the expression levels of RsFT in rootstock plants, alone. The flowering of non-vernalized cabbage would be induced by transmissible agents from rootstocks and not by the expression of cabbage FT, BoFT, from the scion itself.
Jaime A. Teixeira Da Silva; Yaser Hassan Dewir; Adhityo Wicaksono; Leela Sahijram; Haenghoon Kim; Songjun Zeng; Stephen F. Chandler; Munetaka Hosokawa
Folia HorticulturaeDe Gruyter Open Ltd29(2)99 - 1112083-59652017/12[Refereed]
As a result of its domestication, breeding and subsequent commercialization, African violet (Saintpaulia ionantha H. Wendl.) has become the most famous and popular Saintpaulia species. There is interest in producing cultivars that have increased resistance to pests and low temperature, in the introduction of novel horticultural characteristics such as leaf shape, flower colour, size and form, and in improved productivity and enhanced flower duration in planta. In African violet, techniques such as the application of chemical mutagens (ethylmethanesulfonate, N-nitroso-N-methylurea), radiation (gamma (γ)-rays, X-rays, carbon ion beams) and colchicine have been successfully applied to induce mutants. Among these techniques, γ radiation and colchicine have been the most commonly applied mutagens. This review offers a short synthesis of the advances made in African violet breeding, including studies on mutation and somaclonal variation caused by physical and chemical factors, as well as transgenic strategies using Agrobacterium-mediated transformation and particle bombardment. In African violet, Agrobacterium-mediated transformation is affected by the Agrobacterium strain, selection marker, and cutting-induced wounding stress. Somaclonal variation, which arises in tissue cultures, can be problematic in maintaining true-To-Type clonal material, but may be a useful tool for obtaining variation in flower colour. The only transgenic African violet plants generated to date with horticulturally useful traits are tolerant to boron (heavy metal) stress, or bear a glucanase-chitinase gene.
Soo-Jung Yang; Sho Ohno; Ayumi Deguchi; Mitsuru Sato; Mariko Goto; Motoaki Doi; Miki Ohnishi; Fumi Tatsuzawa; Munetaka Hosokawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV22310 - 180304-42382017/09[Refereed]
In this study, we revealed how the petals of Saintpaulia fuse into a corolla by using pinwheel phenotype cultivars. Striped patterns in petal, called pinwheel in Saintpaulia, are attractive phenotypes and thought to be the result of periclinal chimerism. For the selection of a genuine periclinal chimeric cultivar from three pinwheel cultivars, adventitious shoots were induced from leaf lamina. Shoot regeneration was observed from the epidermis in all cultivars by microscopic observation. All regenerated shoots from 'Kaname' flowered as monochromatic pink flowers, corresponding to an Ll phenotype of the cultivar. From the other two cultivars, many shoots flowered not only as an epidermal phenotype but also as a phenotype of the inner layer. In addition, shoot regeneration was induced from epidermis-peeled petioles from these three cultivars. All shoots from 'kaname' flowered as monochromatic blue flowers, corresponding to an L2 phenotype. On the other hand, many shoots from 'Kilauea' flowered not only as monochromatic flowers, corresponding to an L2 phenotype, but also as bi-colored flowers. 'Innocent Pink' did not produce shoots from epidermal-peeled petioles. These results suggested that 'Kaname' is a genuine periclinal chimera, while the other two cultivars have other mechanisms for pinwheel expression. Genomic PCR using primers that amplifies almost the full length of flavonoid 3',5'-hydroxylase (F3'5'H) revealed the gene to be non-functional in pink flowers from Ll of 'Kaname'. From monochromatic pink plants and pink portions of the corolla of 'Raname', full-length F3'5'H was not amplified. Similar results were obtained by quantitative PCR. Finally, we observed the fused portion of the petals and revealed that the petal fusion did not occur by postgenital fusion but by "connection". The process, in Saintpaulia, comprises periclinal cell division in Ll during petal development, active cell division at the edge of the petal, adhesion to the next petals, and fusion. These steps create a striped flower color in Saintpaulia.
Jaime A. Teixeira da Silva; Songjun Zeng; Adhityo Wicaksono; Mafatlal M. Kher; Haenghoon Kim; Munetaka Hosokawa; Yaser Hassan Dewir
SOUTH AFRICAN JOURNAL OF BOTANYELSEVIER SCIENCE BV112501 - 5070254-62992017/09[Refereed]
African violet (Saintpaulia ionantha H. Wendl.) (Gesneriaceae) is a popular ornamental pot plant that is easy to culture ex vitro and in vitro relative to other herbaceous ornamentals. This quality makes it an ideal system for in vitro regeneration experiments. This review summarizes the studies that have been conducted on the in vitro culture and micropropagation of this plant. Although shoot regeneration from internodes, floral buds, anthers and protoplasts has been achieved, leaf blades and petioles have been a popular source tissue for regeneration. An effective and reproducible protocol for the direct induction of shoots, without the formation of any intermediary callus, involves the use of a cytokinin like BA or kinetin, usually in combination with an auxin likeNAA, bothwithin a concentration range of 0.1 to 1 mg/L, and on a Murashige and Skoog basalmedium. Shoots can form in both light and darkness, but most effectively in the light, and rooting can be induced from shoots, even in the absence of an auxin, although a low concentration (0.1-0.5 mg/L) of NAA or IAA is recommended. African violets acclimatize easily, even in a simple soil-based substrate, and can flowerwithin 4 months of transfer to ex vitro conditions. These protocols, including somatic embryogenesis and cryopreservation, may also benefit the in vitro culture and conservation of wild species of Saintpaulia. (C) 2017 SAAB. Published by Elsevier B.V. All rights reserved.
Nao Ota; Tomoyuki Nabeshima; Masahiro Osakabe; Shinichi Aoki; Tatsuya Awano; Munetaka Hosokawa
HORTICULTURE JOURNALJAPAN SOC HORTICULTURAL SCI86(3)349 - 3562189-01022017/07[Refereed]
Although ultraviolet-B (UV-B) light (280-315 nm) irradiation effectively controls spider mites in horticultural crop production, it also causes plant damage, leading to growth suppression, changes in morphology, and leaf scorching. However, sensitivity to UV-B varies among plant species. Here, we assessed the effect of UV-B on growing perilla (Perilla frutescens var. crispa) plants. Three experiments were conducted. In the first experiment, plants of the variety 'Akachirimenshiso' were grown in a plastic house with natural light conditions and subjected to three treatments: natural UV (control), -UV, and natural UV plus artificial UV-B treatment (50 mW.m(-2)) from 0:00 to 3:00. The length of leaf blades that received additional UV-B treatment was reduced by 16.5% and 32.3% in two trials compared with that of the respective controls. In this experiment, additional UV-B irradiation turned the leaf color greener. During UV-B treatment, the values of leaf a*, an indicator of leaf redness, were significantly lower than their respective controls in both trials: 21.5 vs. 31.6 in trial I and 20.2 vs. 30.7 in trial H. For most of the parameters measured in this experiment, no differences were observed between the control and -UV treatment groups. In the second experiment, plants were irradiated with UV-B for 3 weeks at nighttime (0:00-3:00) or daytime (12:00-15:00). In the cultivar 'Houkouakashiso', the length of leaf blades significantly decreased by 15.9% and 20.6% under nighttime UV-B irradiation at 80 and 120 mW.m(2), respectively, compared with that of the non-irradiated controls. Irradiation at 80 mW.m(-2) also decreased the width of the leaf blades by 13.1% and that at 120 mW.m(-2) further decreased it by 25.0%. These results showed that UV-B irradiation at night decreased the size of perilla leaves. In addition, the value of a* became lower under UV-B irradiation in the nighttime. Thus, UV-B irradiation appeared to turn purple perilla leaves green. When plants were irradiated with UV-B in the daytime, there was no significant difference between irradiated and non-irradiated plants in the length or width of leaf blades, a*, or fresh weight of aerial parts and number of nodes on the main stem. In the third experiment, visible rays (VIS) emitted by fluorescent lamps were applied at 0:00-3:00 and 6:00-22:00. Plants were irradiated with 120 mW.m(2) of UV-B at 0:00-3:00. UV-B + VIS treatment of 'Akachirimenshiso' with VIS from fluorescent lamps did not significantly affect the parameters measured in this experiment compared to -UV treatment. The results of this study suggest that UV-B damage to perilla leaves can be avoided by combined irradiation with visible light.
Tomoyuki Nabeshima; Soo-Jung Yang; Sho Ohno; Keita Honda; Ayumi Deguchi; Motoaki Doi; Fumi Tatsuzawa; Munetaka Hosokawa
Frontiers in plant scienceFRONTIERS MEDIA SA81749 - 17491664-462X2017[Refereed]
Saintpaulia ionantha is propagated by adventitious buds in horticulture, and periclinal chimeral cultivars are usually difficult to propagate. However, some periclinal chimeral cultivars can be propagated with adventitious buds, and the mechanism of which has been unknown. Striped flower cultivars "Kaname," "Concord," and "Monique" were used to investigate what causes flower color separation in adventitious shoot-derived plants by tissue culture. These cultivars were revealed to have mutated flavonoid 3', 5' hydroxylase (SiF3'5'H), WDR1 (SiWDR1), or flavonoid 3 hydroxylase (SiF3H), respectively, in their L1 layer. From our previous study using "Kaname," all flowers from adventitious shoots were colored pink, which was the epidermal color of mother plants' flowers. We used "Concrd" and "Monique" from which we obtained not only monochromatic-colored plants the same as the epidermal color of mother plants, but also plants with a monochromatic colored plants, same as the subepidermal color, and a striped flower color the same as mother plants. Histological observations revealed that epidermal cells divided actively at 14 d after culture and they were involved in the formation of adventitious shoots in the cultured leaf segments of "Kaname." On the other hand, in "Concord" and "Monique," the number of divided cells in the subepidermis was rather higher than that of epidermal cells, and subepidermal cells were sometimes involved in shoot formation. In addition, the plant and leaf size of L1-derived plants from "Concord" and "Monique" were non-vigorous and smaller than those derived from the subepidermal layer. In conclusion, periclinal chimeral cultivars of Saintpaulia can be divided into two types. One type has a high cell division activity in the L1 layer, from which only single flower-colored plants derived from L1 can be obtained as adventitious shoots. Another type has a low cell division activity in the L1 layer, from which striped flower-colored plants the same as mother plants derived from several layers including L1 can be obtained as adventitious shoots. In the periclinal chimeral cultivar capable of propagation with adventitious shoots, the possibility was shown that cells in the L2 layer could form shoots by involving cells of the L1 layer with a low division activity.
Tomoyuki Nabeshima; Motoaki Doi; Munetaka Hosokawa
Frontiers in plant scienceFRONTIERS MEDIA SA81940 - 19401664-462X2017[Refereed]
Chrysanthemum stunt viroid (CSVd) was inoculated into two chrysanthemum (Chrysanthemum morifolium) cultivars, the CSVd-susceptible cultivar Piato and the CSVd-resistant cultivar Mari Kazaguruma. For CSVd inoculation, grafting and Agrobacterium-mediated inoculation were used. In grafting experiments, CSVd was detectable in Mari Kazaguruma after grafting onto infected Piato, but after removal of infected rootstocks, CSVd could not be detected in the uppermost leaves. In agroinfection experiments, CSVd systemic infection was observed in Piato but not in Mari Kazaguruma. However, agro-inoculated leaves of Mari Kazaguruma accumulated circular CSVd RNA to levels equivalent to those in Piato at 7 days post-inoculation. In situ detection of CSVd in inoculated leaves revealed that CSVd was absent in phloem of Mari Kazaguruma, while CSVd strongly localized to this site in Piato. We hypothesize that CSVd resistance in Mari Kazaguruma relates not to CSVd replication but to CSVd movement in leaves.
Boubourakas Iraklis; Hiroko Kanda; Tomoyuki Nabeshima; Mayu Onda; Nao Ota; Sota Koeda; Munetaka Hosokawa
Plant cell reportsSPRINGER35(8)1617 - 280721-77142016/08[Refereed]
KEY MESSAGE: CSVd could not infect Nicotiana benthamiana when the plants were pretreated with crude leaf extract of Capsicum chinense 'Sy-2'. C. chinense leaves were revealed to contain strong RNA-digesting activity. Several studies have identified active antiviral and antiviroid agents in plants. Capsicum plants are known to contain antiviral agents, but the mechanism of their activity has not been determined. We aimed to elucidate the mechanism of Capsicum extract's antiviroid activity. Chrysanthemum stunt viroid (CSVd) was inoculated into Nicotiana benthamiana plants before or after treating the plants with a leaf extract of Capsicum chinense 'Sy-2'. CSVd infection was determined using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) 3weeks after inoculation. When Capsicum extract was sprayed or painted onto N. benthamiana before inoculation, it was effective in preventing infection by CSVd. To evaluate CSVd digestion activity in leaf extracts, CSVd was mixed with leaf extracts of Mirabilis, Phytolacca, Pelargonium and Capsicum. CSVd-digesting activities were examined by quantifying undigested CSVd using qRT-PCR, and RNA gel blotting permitted visualization of the digested CSVd. Only Capsicum leaf extract digested CSVd, and in the Capsicum treatment, small digested CSVd products were detected by RNA gel blot analysis. When the digesting experiment was performed for various cultivars and species of Capsicum, only cultivars of C. chinense showed strong CSVd-digesting activity. Our observations indicated that Capsicum extract contains strong RNA-digesting activity, leading to the conclusion that this activity is the main mechanism for protection from infection by CSVd through spraying or painting before inoculation. To our knowledge, this is the first report of a strong RNA-digesting activity by a plant extract.
Li Liu; Jelli Venkatesh; Yeong Deuk Jo; Sota Koeda; Munetaka Hosokawa; Jin-Ho Kang; Sandra Goritschnig; Byoung-Cheorl Kang
TAG. Theoretical and applied genetics. Theoretische und angewandte GenetikSPRINGER129(8)1541 - 560040-57522016/08[Refereed]
KEY MESSAGE: The sy - 2 temperature-sensitive gene from Capsicum chinense was fine mapped to a 138.8-kb region at the distal portion of pepper chromosome 1. Based on expression analyses, two putative F-box genes were identified as sy - 2 candidate genes. Seychelles-2 ('sy-2') is a temperature-sensitive natural mutant of Capsicum chinense, which exhibits an abnormal leaf phenotype when grown at temperatures below 24°C. We previously showed that the sy-2 phenotype is controlled by a single recessive gene, sy-2, located on pepper chromosome 1. In this study, a high-resolution genetic and physical map for the sy-2 locus was constructed using two individual F2 mapping populations derived from a cross between C. chinense mutant 'sy-2' and wild-type 'No. 3341'. The sy-2 gene was fine mapped to a 138.8-kb region between markers SNP 5-5 and SNP 3-8 at the distal portion of chromosome 1, based on comparative genomic analysis and genomic information from pepper. The sy-2 target region was predicted to contain 27 genes. Expression analysis of these predicted genes showed a differential expression pattern for ORF10 and ORF20 between mutant and wild-type plants; with both having significantly lower expression in 'sy-2' than in wild-type plants. In addition, the coding sequences of both ORF10 and ORF20 contained single nucleotide polymorphisms (SNPs) causing amino acid changes, which may have important functional consequences. ORF10 and ORF20 are predicted to encode F-box proteins, which are components of the SCF complex. Based on the differential expression pattern and the presence of nonsynonymous SNPs, we suggest that these two putative F-box genes are most likely responsible for the temperature-sensitive phenotypes in pepper. Further investigation of these genes may enable a better understanding of the molecular mechanisms of low temperature sensitivity in plants.
Tomoyuki Nabeshima; Motoaki Doi; Munetaka Hosokawa
Journal of virological methodsELSEVIER SCIENCE BV234169 - 730166-09342016/08[Refereed]
Agroinfiltration was tested as a method of inoculation of chrysanthemum plants with chrysanthemum stunt viroid (CSVd). Binary vectors harboring dimeric CSVd sequences in sense and antisense orientations were constructed, and Agrobacterium transfected with these binary vectors was infiltrated into chrysanthemum leaves. Northern blotting and reverse transcription polymerase chain reaction analysis showed that local infection was established within 7 days and systemic infection within 20 days. CSVd polarities showed no difference in infectivity. This study showed that agroinfiltration of chrysanthemum plants is an easy, rapid, and cost-effective method for CSVd inoculation.
Sho Ohno; Wakako Hori; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
HORTICULTURE JOURNALJAPAN SOC HORTICULTURAL SCI85(2)177 - 1862189-01022016/04[Refereed]
Bicolor flowering dahlias generally produce inflorescences with bicolor petals characterized by a colored basal part and a white tip; however, they frequently produce single-colored petals. This petal color lability prevents uniform production of cut or pot flowers of bicolor dahlias and reduces the economic value of bicolor cultivars. In this study, to reveal the underlying mechanism and control color lability, the pattern of occurrence of single colored petals was characterized in a red white bicolor flowering cultivar 'Yuino'. 'Yuino' produced inflorescences with bicolor petals, red petals, and both red and bicolor petals. Red petals occurred almost always at the outer whorls or sectorally in a mixed inflorescence, similar to a chimera or a lateral mutant. The occurrence of red petals was higher in field experiments during May to December than in greenhouse experiments during October to next July. We identified the "R-line" plant, which produced red petals with high frequency during the winter to spring cultivation; this characteristic to produce red petals with high frequency was retained through vegetative propagation. There were strong relationships between inflorescence color and leaf phenotype; red petal-producing plants accumulated flavonoids in leaves, whereas only bicolor petal-producing plants tended not to accumulate flavonoid in leaves. This suggests that petal color of 'Yuino' is associated with flavonoid synthetic potential in shoot. Therefore, a phenotypic difference is observed not only in petal colors but also at the whole plant level.
Fumi Tatsuzawa; Munetaka Hosokawa
HORTICULTURE JOURNALJAPAN SOC HORTICULTURAL SCI85(1)63 - 692189-01022016/01[Refereed]
The flower colors and anthocyanin constitution of sixteen cultivars of Saintpaulia were surveyed to determine the relationship between their flower colors and anthocyanin components. Six anthocyanins were isolated from the flowers of these cultivars as major anthocyanins along with three minor ones, and their structures were identified by co-HPLC or chemical and spectroscopic techniques. Among them, a novel anthocyanin, pelargonidin 3-O-[6-O-(4-O-(acetyl)-alpha-rhamnopyranosyl)-beta-glucopyranoside] (pelargonidin 3-acetyl-rutinoside; 8) was found in cultivars of 'Georgia' and 'Jessica' as a major anthocyanin. Regarding the flower color variation in these cultivars, the hue values (b*/a*) of these flower colors were responsible for the glycosidic positions in the anthocyanidin molecule and also the combination of anthocyanins. These flower colors were classified into six groups, A-F, based on the flower colors and anthocyanin components were arranged as follows. In violet-blue flowers of group A (hue values b*/a* = -2.61--1.72, VB N89B-VB 94B) and purple-violet flowers of group B (-1.06 and -0.81, PV N82A and PV N80B), malvidin 3-acetyl-rutinoside-5-glucoside was the most effective major anthocyanin for flower colors. In purple-violet flowers (-0.69 and -0.53, PV N80B and PV N81A) of group C, peonidin 3-acetyl-rutinoside-5-glucoside was the most effective major anthocyanin for flower colors. In red-purple flowers (-0.44--0.27, RP 73A-RP N74B) of groups D, pelargonidin 3-acetyl-rutinoside-5-glucoside, in red-purple flowers (-0.03 and -0.02, RP 60D and RP 71D) of group E, pelargonidin 3-acetyl-rutinoside, and in red-purple flowers (0.04 and 0.13, RP 61A and RP 71A) of group F, peonidin 3-acetyl-rutinoside were the most effective major anthocyanins for flower colors. From these results, the glucosylation of 5-OH in anthocyanidin 3-acetyl-rutinoside and an increase in the methylation of the B-ring in anthocyanidin were considered to have the most important effects on flower color variations in these Saintpaulia cultivars.
Deguchi Ayumi; Tatsuzawa Fumi; Hosokawa Munetaka; Doi Motoaki; Ohno Sho
The Horticulture JournalThe Japanese Society for Horticultural Science85(4)340 - 3502189-01022016
The black flower color of dahlias (Dahlia variabilis) has been suggested to be attributed to a high accumulation of cyanidin (Cy)-based anthocyanins. A possible explanation for this effect is that Cy-based anthocyanins in dahlias contribute more to the black flower color than pelargonidin (Pg)-based anthocyanins by lowering petal lightness (L*) and chroma (C*), but no obvious evidence has been reported. In this study, four major anthocyanins accumulated in dahlia petals, 3,5-diglucoside (3,5diG) and 3-(6''-malonylglucoside)-5-glucoside (3MG5G) of Pg and Cy, were purified and their colors were evaluated in vitro at various pHs (3.0, 4.0, 4.5, 5.0, 5.5, 6.0, or 7.0) and various concentrations (0.25, 0.5, 1.0, 2.0, or 3.0mg·mL−1 at pH5.0 or pH3.0). The color of solution of purified anthocyanins varied depending on pH. At pH5.0, which is approximately the same as pH of dahlia petals, and at pH3.0, at which anthocyanins are relatively stable, the L* and C* of Cy 3,5diG were similar to or higher than those of Pg 3,5diG, suggesting that Cy 3,5diG did not contribute more to the black flower coloring than Pg 3,5diG. On the other hand, the L* and C* of Cy 3MG5G were significantly lower than those of Pg 3MG5G, particularly above 2.0mg·mL−1, suggesting that Cy 3MG5G contributed more than Pg 3MG5G. A similar tendency was observed in the color measurement of mixed anthocyanins in various proportion of Pg and Cy. The L* and C* of Pg 3MG5G were much higher than those of the other three anthocyanins; therefore, its color was considered to be the farthest from black among the four anthocyanins. The accumulated amount of 3MG5G-type anthocyanins was much higher than that of 3,5diG-type anthocyanins in all nine cultivars, although the proportion of Pg- and Cy-based anthocyanins varied among the cultivars. Considering these results, it was suggested that because 3MG5G-type anthocyanins predominantly accumulate in petals, and Cy 3MG5G has a significantly higher contribution to lowering L* and C* than Pg 3MG5G, the high accumulation of Cy-based anthocyanins is critical for the black flower coloring of dahlias. The contribution of each anthocyanin is considered to depend on the structure; therefore, identifying the anthocyanin with the highest contribution to lowering L* and C* may enable the production of black flowers in various species through the high accumulation of the anthocyanin in petals.
Sota Koeda; Elly Kesumawati; Yuri Tanaka; Munetaka Hosokawa; Motoaki Doi; Akira Kitajima
Tropical Agriculture and DevelopmentJapanese Society for Tropical Agriculture60(2)64 - 641882-84692016[Refereed]
Hot peppers (Capsicum spp.) are essential spice in Indonesian meal. Recently, diseases caused by viruses are emerging as serious problems in pepper production at northern Sumatra. Viruses are destabilizing farmers' incomes and the prices of spice used daily by the consumers. In the present study, to obtain the fundamental knowledge of severe damage in pepper production, field research and molecular study were conducted to assess the genetic identity and diversity of pepper-infecting viruses. Five local farmers' fields located at the suburbs of Banda Aceh were chosen for the study. At each field approximately 500 to 3,000 plants of C. annuum were cultivated. The yellow leaf curl symptoms were observed in more than 81 % of plants in all the fields, and symptoms reached 100 % at four fields out of five. As yellow leaf curl symptoms are often caused by begomoviruses, DNA-A full-length sequences were determined and clarified that Pepper yellow leaf curl Indonesia virus (PepYLCIV), Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), and Ageratum yellow vein virus (AYVV) were infecting pepper plants. By detecting three different begomoviruses independently using specific primers, it was suggested that most of pepper plants were infected with two to three viruses. The present study inferred that mixed infection of begomoviruses is associated with the serious virus problems in pepper production at northern Sumatra, Indonesia.
Ayumi Deguchi; Fumi Tatsuzawa; Munetaka Hosokawa; Motoaki Doi; Sho Ohno
PlantaSPRINGER242(3)663 - 750032-09352015/09[Refereed]
MAIN CONCLUSION: Tobacco streak virus suppressed post-transcriptional gene silencing and caused a flower color change in black dahlias, which supported the role of cyanidin-based anthocyanins for black flower appearance. Black flower color of dahlia (Dahlia variabilis) has been attributed, in part, to the high accumulation of cyanidin-based anthocyanins that occurs when flavone synthesis is reduced because of post-transcriptional genesilencing (PTGS) of flavone synthase II (DvFNS). There are also purple-flowering plants that have emerged from a black cultivar 'Kokucho'. We report that the purple color is not caused by a mutation, as previously thought, but by infection with tobacco streak virus (TSVdahlia), which suppresses the PTGS of DvFNS. When TSVdahlia was eliminated from the purple-flowering 'Kokucho' by leaf primordia-free shoot apical meristem culture, the resulting flowers were black. TSVdahlia-infected purple flowers had lower numbers of siRNAs to DvFNS than black flowers, suggesting that TSVdahlia has a silencing suppressor. The graft inoculation of other black cultivars with TSVdahlia altered their flower color drastically except for 'Fidalgo Blacky', a very deep black cultivar with the highest amount of cyanidin-based anthocyanins. The flowers of all six TSVdahlia-infected cultivars accumulated increased amounts of flavones and reduced amounts of cyanidin-based anthocyanins. 'Fidalgo Blacky' remained black despite the change in pigment accumulation, and the amounts of cyanidin-based anthocyanins in its TSVdahlia-infected plants were still higher than those of other cultivars. We propose that black flower color in dahlia is controlled by two different mechanisms that increase the amount of cyanidin-based anthocyanins: DvFNS PTGS-dependent and -independent mechanisms. If both mechanisms occur simultaneously, the flower color will be blacker than if only a single mechanism is active.
Mahmoud Sharaf-Eldin; Shereen Elkholy; Munetaka Hosokawa; Kai Yanagawa; Eiji Nawata; Keiichi Takagi; Jose-Antonio Fernandez
ZEITSCHRIFT FUR ARZNEI- & GEWURZPFLANZENAGRIMEDIA GMBH20(2)84 - 871431-92922015/06[Refereed]
The detection of new mutations in saffron flowers with increased number of stigmata has been reported, and the other mutants in floral whorls were discussed. Referable to the sexual sterility of saffron, it is concluded that if the observed mutations verified to be external stimuli determined; the cause which may be detected could increase saffron spice production and; reduce the cost of saffron production in the future. Mutagenic effects of carbon-ion beam on corms of saffron have been reported in this study.
Fumi Tatsuzawa; Sayumi Matsuda; Kazuhisa Kato; Munetaka Hosokawa
HORTICULTURE JOURNALJAPAN SOC HORTICULTURAL SCI84(1)77 - 822189-01022015/01[Refereed]
A new acetylated anthocyanin was extracted from the red-purple flowers of Saintpaulia 'Tomoko' with 5% HOAc-H2O or 5% formic acid-H2O, and determined to be peonidin 3-O-[6-O-(4-O-(acetyl)-alpha-rhamnopyranosyl)-beta-glucospyranoside] (1), by chemical and spectroscopic methods. In addition, two known acetylated cyanidin glycosides, cyanidin 3-O-[6-O-(4-O-(acetyl)-alpha-rhamnopyranosyl)-beta-glucospyranoside] (2) and cyanidin 3-O-[2-O-(beta-xylopyranosyl)-6-O-(acetyl)-beta-glucopyranoside] (3), were also identified in the redpurple flowers and grayed-purple leaves of S. 'Tomoko', respectively. These three anthocyanins have not been reported hitherto in plant tissues in the genus Saintpaulia.
Sayumi Matsuda; Mitsuru Sato; Sho Ohno; Soo-Jung Yang; Motoaki Doi; Munetaka Hosokawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI83(4)308 - 3161882-33512014/10[Refereed]
For determination of the endogenous and exogenous causes of somaclonal variation in in vitro culture, a bioassay system was developed using the variegated Sairupaulia (African violet) 'Thamires' (Saintpaulia sp.), having pink petals with blue splotches caused by transposon VGs1 (Variation Generator of Saintpaulia 1) deletion in the promoter region of flavonoid 3',5'-hydroxylase. Not only true-to-type but also many solid blue and chimeric plants regenerate in vitro-cultured explants of this cultivar. Using multiplex PCR that enables the determination of these variations, we attempted to evaluate the effects of four candidate triggers of mutation: pre-existing mutated cells, shooting conditions in vitro or ex vitro, cutting treatment of explants, and addition of plant growth regulators (PGRs) to the medium. The percentages of somaclonal variations among total shoots regenerated from leaf segments and stamens were 46.6 and 56.5, which were higher than the percentages expected from pre-existing mutated cells (3.6 and 1.4, respectively). These results indicate that pre-existing mutated cells are not a main cause of somaclonal variations. The percentage of somaclonal variation was independent of culture conditions for mother plants; the mutation percentages of adventitious shoots regenerated from ex vitro- and in vitro-grown leaves were 9.2% and 8.5%, respectively. In addition, the percentage of somaclonal variations of adventitious shoots regenerated under in vitro conditions from the in vitro grown mother plants was also low, at 4.9%. This indicates that the in vitro condition itself is not a main cause of somaclonal variation. However, when adventitious shoots were regenerated from 10 x 5-mm cut-leaf laminas on a PGR-free medium, the percentage of somaclonal variation was 26.4%. In addition, the percentage of somaclonal variations dramatically increased when PGRs were added to the medium for both leaves and leaf segments (39.9 and 46.6, respectively). The bioassay system using Saintpaulia 'Thamires' will enable the screening of many environmental factors because of its rapidity and ease of use and will facilitate the development of a new tissue culture technology for avoiding mutation.
Sota Koeda; Kosuke Sato; Kenichi Tomi; Yoshiyuki Tanaka; Rihito Takisawa; Munetaka Hosokawa; Motoaki Doi; Tetsuya Nakazaki; Akira Kitajima
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI83(3)244 - 2511882-33512014/07[Refereed]
'No.80' (Capsicum chinense) from the Caribbean is a valuable genetic source from the aspect of its non-pungent and highly aromatic traits. In the present study, the non-pungency, volatile components, and phylogenetic origin of 'No.80' were analyzed with another C. chinense cultivar, 'No.2' from Brazil, which is also non-pungent but less aromatic. Expressions and deduced amino acid sequences of acyltransferase (Punl) of 'No.80' and 'No.2' were normal compared with a pungent cultivar, 'Habanero'. Insertions of 7-bp and 8-bp resulting in frameshift mutations were found in the coding regions of putative aminotransferase (p-AMT) of 'No.80' and 'No.2', respectively. Co-segregation of these insertions with the non-pungent phenotypes in F-1 and F-2 populations obtained from crossing 'No.80' or 'No.2' with 'Habanero' suggested that non-pungency in these cultivars arose from genetic mutations of p-AMT that occurred independently. Moreover, molecular phylogenetic analysis suggested that 'No.80', a close relative of 'No.2', originates from capsic*ms migrated from the South American mainland. In addition to pungency, we assessed the volatile components of the highly aromatic 'No.80', the less aromatic 'No.2', and their F-1 hybrid using gas chromatography. 'No.80' contained higher levels of aroma-contributing volatiles than 'No.2', which correlated with the stronger and weaker aromas of two cultivars. Further, the fruit of F-1 progenies emitted a number of volatile compounds between or higher than their corresponding parents. Based on these results, the approaches for breeding highly aromatic non-pungent cultivars are discussed.
T. Nabeshima; M. Hosokawa; S. Yano; K. Ohishi; M. Doi
JOURNAL OF HORTICULTURAL SCIENCE & BIOTECHNOLOGYHEADLEY BROTHERS LTD89(1)29 - 341462-03162014/01[Refereed]
A total of 199 chrysanthemum (Chrysanthemum x morifolium Ramat.) cultivars were used in screening for resistance to chrysanthemum stunt viroid (CSVd). Plants were inoculated with CSVd by attaching leaf primordia-free, shoot apical meristems to CSVd-infected root tips of a CSVd-sensitive cultivar. CSVd titres in the first and third youngest expanded leaves were analysed after 2 months in culture, and a systematic classification into four groups was conducted. Long-term cultivation records provided by a breeding nursery confirmed our findings of those screened cultivars classified in vitro as "non-detected-type" (i.e., those expected to have a high level of resistance). In addition, of the 15 candidate cultivars selected as "non-detected-type", four cultivars were shown to have a high level of CSVd-resistance by ex vitro grafting experiments. From the results of our in vitro and ex vitro experiments, the advantages of our newly-developed in vitro-screening system are discussed.
Yoshiyuki Tanaka; Hirotsugu Yoneda; Munetaka Hosokawa; Tetsuya Miwa; Susumu Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV165242 - 2450304-42382014/01[Refereed]
Capsinoids are a group of low-pungent capsaicinoid analogs produced in Capsicum fruits. Their bioactivities are similar to those of capsaicinoids, such as the suppression of fat accumulation and antioxidant activity. Capsinoids have low pungency and thus are more easily used as food ingredients than capsaicinoids. Capsinoids are, however, unstable in water and upon heating, and their content decreases during fruit maturation. Therefore, capsinoid-containing fruits should be consumed raw and before maturation. Previous genetic study showed that capsinoid biosynthesis is controlled by two genes, p-AMT and Pun1, designated as A and B, respectively. It is expected that aaBB and aaBb represent low-pungent plants containing capsinoids. In this study, DNA markers of p-AMT and Pun1 were used in an attempt to develop a new fresh cultivar containing capsinoids in a cross-breeding program. The genotypes of each progeny derived from the cross of 'Murasaki' (AAbb) x 'CH-19 Sweet' (aaBB) were determined using the DNA markers. Specifically, p-AMT genotypes were determined using a dCAPS marker and Pun1 genotypes using a SCAR marker. From the results of genotyping, aaBB or aaBb plants were selected and developed as a new cultivar, 'Maru Salad'. The capsinoid content of fruits of 'Maru Salad' at harvest was approximately 700 mu g/gDW. Fruits of 'Maru Salad' are larger than those of 'CH-19 Sweet' and are suitable to be consumed raw. (C) 2013 Elsevier B.V. All rights reserved.
Sota Koeda; Munetaka Hosokawa; Hiroki Saito; Motoaki Doi
Journal of plant researchSPRINGER JAPAN KK126(5)675 - 840918-94402013/09[Refereed]
Plants in tropical regions experience temperature fluctuation only in non-extreme ambient temperatures. Thus, moderate changes in temperatures, which they never experience in their local environments, might be sufficient to manifest the locally hidden phenotype caused by natural mutation. To validate this hypothesis, temperature-treating experiments were performed on Capsicum accessions collected from tropical regions. Thirty-six Capsicum accessions, collected from Caribbean countries, were screened for temperature sensitivity. Similarities in their temperature sensitivities were compared with Sy-2 (C. chinense) from Seychelles, which was previously found to be a temperature-sensitive accession. Tr-13 from Trinidad & Tobago exhibited developmental abnormalities at temperatures below 24°C. Expression of defense-related genes was induced, and salicylic acid, which is a key molecule in the plant's defense response, accumulated in Tr-13 at temperatures below 24°C. Tr-13 and Sy-2 appeared normal when they were grown at temperatures simulating those in Trinidad and Seychelles, respectively. Crossing Tr-13 with No. 3341 or Sy-2 revealed that the temperature-sensitive phenotype of Tr-13 was caused by a genetic mutation in the same locus as Sy-2. Plants having a temperature-sensitive phenotype that is caused by natural mutations evade artificial selection and exist as crops in specific environments, such as tropical regions.
Sho Ohno; Ayumi Deguchi; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
PlantaSPRINGER238(2)331 - 430032-09352013/08[Refereed]
The study was aimed to identify the factors that regulate the intensity of flower color in cyanic dahlia (Dahlia variabilis), using fifteen cultivars with different color intensities in their petals. The cultivars were classified into three groups based on their flavonoid composition: ivory white cultivars with flavones; purple and pink cultivars with flavones and anthocyanins; and red cultivars with flavones, anthocyanins, and chalcones. Among the purple, pink, and ivory white cultivars, an inverse relationship was detected between lightness, which was used as an indicator for color intensity and anthocyanin content. A positive correlation was detected between anthocyanin contents and the expression of some structural genes in the anthocyanin synthesis pathway that are regulated by DvIVS, a basic helix-loop-helix transcription factor. A positive correlation between anthocyanin content and expression of DvIVS was also found. The promoter region of DvIVS was classified into three types, with cultivars carrying Type 1 promoter exhibited deep coloring, those carrying Type 2 and/or Type 3 exhibited pale coloring, and those carrying Type 1 and Type 2 and/or Type 3 exhibited medium coloring. The transcripts of the genes from these promoters encoded full-length predicted proteins. These results suggested that the genotype of the promoter region in DvIVS is one of the key factors determining the flower color intensity.
Ayumi Deguchi; Sho Ohno; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
PlantaSPRINGER237(5)1325 - 350032-09352013/05[Refereed]
Black color in flowers is a highly attractive trait in the floricultural industry, but its underlying mechanisms are largely unknown. This study was performed to identify the bases of the high accumulation of anthocyanidins in black cultivars and to determine whether the high accumulation of total anthocyanidins alone leads to the black appearance. Our approach was to compare black dahlia (Dahlia variabilis) cultivars with purple cultivars and a purple flowering mutant of a black cultivar, using pigment and molecular analyses. Black cultivars characteristically exhibited low lightness, high petal accumulation of cyanidin and total anthocyanidins without flavones, and marked suppression of flavone synthase (DvFNS) expression. A comparative study using black and purple cultivars revealed that neither the absence of flavones nor high accumulation of total anthocyanidins is solely sufficient for black appearance, but that cyanidin content in petals is also an important factor in the phenotype. A study comparing the black cultivar 'Kokucho' and its purple mutant showed that suppression of DvFNS abolishes the competition between anthocyanidin and flavone synthesis and leads to accumulation of cyanidin and total anthocyanidins that produce a black appearance. Surprisingly, in black cultivars the suppression of DvFNS occurred in a post-transcriptional manner, as determined by small RNA mapping.
Munetaka Hosokawa; Takayoshi Yamauchi; Masayoshi Takahama; Mariko Goto; Sachiko Mikano; Yuki Yamaguchi; Yoshiyuki Tanaka; Sho Ohno; Sota Koeda; Motoaki Doi; Susumu Yazawa
Plant cell reportsSPRINGER32(5)601 - 90721-77142013/05[Refereed]
KEY MESSAGE: The corolla of Petunia 'Magic Samba' exhibits unstable anthocyanin expression depending on its phosphorus content. Phosphorus deficiency enhanced post-transcriptional gene silencing of chalcone synthase - A in the corolla. Petunia (Petunia hybrida) 'Magic Samba' has unstable red-white bicolored corollas that respond to nutrient deficiency. We grew this cultivar hydroponically using solutions that lacked one or several nutrients to identify the specific nutrient related to anthocyanin expression in corolla. The white area of the corolla widened under phosphorus (P)-deficient conditions. When the P content of the corolla grown under P-deficient conditions dropped to <2,000 ppm, completely white corollas continued to develop in >40 corollas until the plants died. Other elemental deficiencies had no clear effects on anthocyanin suppression in the corolla. After phosphate was resupplied to the P-deficient plants, anthocyanin was restored in the corollas. The expression of chalcone synthase-A (CHS-A) was suppressed in the white area that widened under P-suppressed conditions, whereas the expression of several other genes related to anthocyanin biosynthesis was enhanced more in the white area than in the red area. Reddish leaves and sepals developed under the P-deficient condition, which is a typical P-deficiency symptom. Two genes related to anthocyanin biosynthesis were enhanced in the reddish organs. Small interfering RNA analysis of CHS-A showed that the suppression resulted from post-transcriptional gene silencing (PTGS). Thus, it was hypothesized that the enhancement of anthocyanin biosynthetic gene expression due to P-deficiency triggered PTGS of CHS-A, which resulted in white corolla development.
Motoaki Doi; Nozomi Nakamura; Youko Takizawa; Makiko Wakita; Eumiya Shimizu; Yoshikuni Kitamura; Munetaka Hosokawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV150441 - 4470304-42382013/02[Refereed]
The harvest characteristics of canna (Canna x generalis L.H. Bailey) 'Butter Cup' leaves were investigated. Leaves harvested during the daytime under fair conditions inrolled rapidly and then browned. The severities of these physiological disorders were closely related to the leaf age and harvest time. Aged leaves harvested from shoots with senescing inflorescences exhibited severe disorders, whereas young leaves harvested from shoots with tight inflorescences exhibited less physiological disorders. In addition, aged leaves harvested just before sunrise did not show any physiological disorders, but the severity of leaf inrolling and browning increased with the increase in integrated solar radiation at harvest. The transpiration rate of young leaves decreased after harvest, whereas that of aged leaves sharply increased 10 min after harvest and then gradually decreased. The leaf resistance of both young and aged leaves was inversely correlated to transpiration rates. The behavior of stomata after harvest was also consistent with these two parameters, suggesting that loss of water is responsible for inrolling of aged leaves. Three min after harvest, a rapid change in the leaf electric potential was detected in aged leaves. Several postharvest treatments indicated that suppressing transpiration rates inhibit both leaf inrolling and leaf browning, and that oxygen is needed for leaf browning. However, none of the treatments were effective in preventing the leaf inrolling and browning after the treatments were conducted and leaves were exposed to ambient air. (C) 2012 Elsevier B.V. All rights reserved.
Three acylated anthocyanins and a flavone glycoside in violet-blue flowers of Saintpaulia 'Thamires'
F. Tatsuzawa; M. Hosokawa; N. Saito; T. Honda
SOUTH AFRICAN JOURNAL OF BOTANYELSEVIER SCIENCE BV7971 - 760254-62992012/03[Refereed]
Three new acylated anthocyanidin 3-rutinoside-5-glucosides were isolated from the violet-blue flowers of Saintpaulia 'Thamires' (Saintpaulia sp.) along with a known flavone glycoside. Three new acetylated anthocyanins were determined to be 3-O-[6-O-(4-O-(acetyl)-alpha-rhamnopyranosyl)-beta-glucopyranoside]-5-O-(beta-glucopyranoside)s of malvidin (pigment 1), peonidin (pigment 2), and pelargonidin (pigment 3) by chemical and spectroscopic methods. HPLC analysis revealed that malvidin 3-O-acetylrutinoside-5-O-glucoside existed as a dominant pigment in the violet-blue flowers. Moreover, the isolated flavone was identified to be apigenin 4'-O-beta-glucuronopyranoside (pigment 4).On the visible absorption spectral curves of fresh violet-blue petals and in their crude extracts in pH 5.0 buffer solution, two characteristic absorption maxima at 547 and 577 nm, with a shoulder near 620 nm, were observed. In contrast, the absorption curves of malvidin 3-O-acetylrutinoside-5-O-glucoside and its deacyl anthocyanin exhibited only one maximum at 535 nm in pH 5.0 buffer solution, and its color was violet and soon fell into decay.However, by addition of apigenin 4'-O-glucuronide, the color of malvidin 3-O-acetylrutinoside-5-O-glucoside changed from violet to violet-blue, similar to that of the fresh flower in pH 5.0 buffer solution. The absorption curve of its violet-blue solution exhibited two similar absorption maxima at 547 and 577 nm, with a shoulder near 620 nm. These results suggest that intermolecular copigmentation between malvidin 3-O-acetylrutinoside-5-O-glucoside and apigenin 4'-O-glucuronide may be responsible for the violet-blue flower color of S. 'Thamires'. (C) 2011 SAAB. Published by Elsevier B.V. All rights reserved.
Nabeshima Tomoyuki; Hosokawa Munetaka; Yano Shinobu; Ohishi Kazushi; Doi Motoaki
Journal of the Japanese Society for Horticultural ScienceTHE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCE81(3)285 - 2941882-33512012
Cultivars resistant to chrysanthemum stunt viroid (CSVd) are desirable for stable production of chrysanthemum (Chrysanthemum × morifolium Ramat.). We previously reported that CSVd was absent not only from shoot apical meristems (SAMs) but also from leaf primordia (LP) of resistant plants following CSVd inoculation. Using this characteristic as a phenotypic marker, we could identify several resistant cultivars in this study. CSVd was inoculated directly into the SAMs of 85 commercial cultivars by attaching SAMs to infected root tips in vitro, and the cultivars were tested for the presence of CSVd in newly expanded leaves. Of the 85 cultivars, 20 resistant candidates could be identified. These plants were classified into two types according to their CSVd infection characteristics: a CSVd-uninfected or slow titer-increasing type and a CSVd-disappearance type. Scions of four of the 20 candidate cultivars were grafted to CSVd-infected rootstocks for evaluation of the CSVd titer in newly expanded leaves. Although CSVd was detected at high titer in two candidate cultivars over the two-month testing period, ‘Sei no Issei’ and ‘Mari Kazaguruma’, other two candidate cultivars, proved to be resistant cultivars. In ‘Sei no Issei’, CSVd was detected temporarily after grafting, but titers decreased in the newly expanded leaves, while CSVd was not detected in a shoot tip of an infected plant and only at trace levels in a young leaf. ‘Mari Kazaguruma’ also proved to be a resistant cultivar with a slow titer increase. The different types of CSVd resistance in chrysanthemum cultivars will contribute to CSVd-resistant breeding. With some modifications, the in vitro screening method established here will be available for broadening the diversity of genetic resources resistant to CSVd.
Sota Koeda; Munetaka Hosokawa; Byoung-Cheorl Kang; Chihiro Tanaka; Doil Choi; Satoshi Sano; Takashi Shiina; Motoaki Doi; Susumu Yazawa
Journal of plant researchSPRINGER TOKYO125(1)137 - 450918-94402012/01[Refereed]
Temperature is one of the most important environmental factors that influence plant growth and development. Recent studies imply that plants show various responses to non-extreme ambient temperatures. Previously, we have found that a pepper cultivar cv. Sy-2 (Capsicum chinense) shows developmental defects at temperatures below 24°C. In this study, to gain new insights into the temperature sensitivity of cv. Sy-2, temperature-sensitive genes were screened using microarray techniques. At restrictive temperature of 20°C, almost one-fourth of the 411 up-regulated genes were defense related or predicted to be defense related. Further expression analyses of several defense-related genes showed that defense-related genes in cv. Sy-2 were constitutively expressed at temperatures below 24°C. Moreover, accumulation of high level of salicylic acid (SA) in cv. Sy-2 grown at 20°C suggests that the defense response is activated in the absence of pathogens. To confirm that the defense response is induced in cv. Sy-2 below 24°C, we evaluated the resistance to biotrophic bacterial pathogen Xanthom*onas campestris pv. vesicatoria and necrotrophic fungal pathogen Cercospora capsici. Cv. Sy-2 showed enhanced resistance to X. campestris pv. vesicatoria, but not to C. capsici.
Sho Ohno; Munetaka Hosokawa; Misa Kojima; Yoshikuni Kitamura; Atsushi Hoshino; Fumi Tatsuzawa; Motoaki Doi; Susumu Yazawa
PlantaSPRINGER234(5)945 - 580032-09352011/11[Refereed]
Garden dahlias (Dahlia variabilis) are autoallooctoploids with redundant genes producing wide color variations in flowers. There are no pure white dahlia cultivars, despite its long breeding history. However, the white areas of bicolor flower petals appear to be pure white. The objective of this experiment was to elucidate the mechanism by which the pure white color is expressed in the petals of some bicolor cultivars. A pigment analysis showed that no flavonoid derivatives were detected in the white areas of petals in a star-type cultivar 'Yuino' and the two seedling cultivars 'OriW1' and 'OriW2' borne from a red-white bicolor cultivar, 'Orihime', indicating that their white areas are pure white. Semi-quantitative RT-PCR showed that in the pure white areas, transcripts of two chalcone synthases (CHS), DvCHS1 and DvCHS2 which share 69% nucleotide similarity with each other, were barely detected. Premature mRNA of DvCHS1 and DvCHS2 were detected, indicating that these two CHS genes are silenced post-transcriptionally. RNA gel blot analysis revealed that small interfering RNAs (siRNAs) derived from CHSs were produced in these pure white areas. By high-throughput sequence analysis of small RNAs in the pure white areas with no mismatch acceptance, small RNAs were mapped to two alleles of DvCHS1 and two alleles of DvCHS2 expressed in 'Yuino' petals. Therefore, we concluded that simultaneous siRNA-mediated post-transcriptional gene silencing of redundant CHS genes results in the appearance of pure white color in dahlias.
Sho Ohno; Munetaka Hosokawa; Atsushi Hoshino; Yoshikuni Kitamura; Yasumasa Morita; Kyeung-Ii Park; Akiko Nakashima; Ayumi Deguchi; Fumi Tatsuzawa; Motoaki Doi; Shigeru Iida; Susumu Yazawa
Journal of experimental botanyOXFORD UNIV PRESS62(14)5105 - 160022-09572011/10[Refereed]
Dahlias (Dahlia variabilis) exhibit a wide range of flower colours because of accumulation of anthocyanin and other flavonoids in their ray florets. Two lateral mutants were used that spontaneously occurred in 'Michael J' (MJW) which has yellow ray florets with orange variegation. MJOr, a bud mutant producing completely orange ray florets, accumulates anthocyanins, flavones, and butein, and MJY, another mutant producing completely yellow ray florets, accumulates flavones and butein. Reverse transcription-PCR analysis showed that expression of chalcone synthase 1 (DvCHS1), flavanone 3-hydroxylase (DvF3H), dihydroflavonol 4-reductase (DvDFR), anthocyanidin synthase (DvANS), and DvIVS encoding a basic helix-loop-helix transcription factor were suppressed, whereas that of chalcone isomerase (DvCHI) and DvCHS2, another CHS with 69% nucleotide identity with DvCHS1, was not suppressed in the yellow ray florets of MJY. A 5.4 kb CACTA superfamily transposable element, transposable element of Dahlia variabilis 1 (Tdv1), was found in the fourth intron of the DvIVS gene of MJW and MJY, and footprints of Tdv1 were detected in the variegated flowers of MJW. It is shown that only one type of DvIVS gene was expressed in MJOr, whereas these plants are likely to have three types of the DvIVS gene. On the basis of these results, the mechanism regulating the formation of orange and yellow ray florets in dahlia is discussed.
Munetaka Hosokawa; Hisanao Suzue; Takashi Fudano; Motoaki Doi
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI80(4)461 - 4681882-33512011/10[Refereed]
Vigorous shoots in chrysanthemums (Chrysanthemum morifolium) often develop from shoot tips (apical meristems with two leaf primordia) cultured in vitro following particle bombardment. The average fresh weight of regenerated shoots from a bombarded shoot tip of chrysanthemum 'Jinba' was 10 times more than that regenerated from unbombarded shoot tips. The average number of leaves per bombarded shoot tip was also more than that from an unbombarded shoot tip. The average number of leaves developing from a shoot tip increased with an increase in the amount of gold particles shot into the shoot tips. In addition, when the area destroyed in a shoot apical meristem (SAM) was varied by bombardment through nylon mesh with different pore sizes, the total number of leaves produced from each shoot tip increased with the size of the destroyed SAM area. Knowing the origin of these vigorous shoots, which may be from the bombarded meristem or from the lateral meristems, is important for the screening of transgenic plants. When the entire surface of a SAM was destroyed by bombardment, it was unable to rebuild itself; instead, lateral meristems were initiated at the base of the leaf primordia. Furthermore, the initiation of lateral meristems at the base of the leaf primordia was also observed in instances of restoration of the area of a partially destroyed SAM. This result indicates that vigorous lateral shoots initiate and develop from the bases of leaf primordia when SAMs are damaged to varying degrees. When leaf primordia-free shoot apical meristems (LP-free SAMs) were cultured after bombardment, vigorous shoots failed to develop from the wounded SAM; instead, the wounded LP-free SAMs regenerated a SAM by repairing the wounded areas and developed a non-vigorous single shoot. It was concluded the vigorous shoots do not participate in transgenic plant production because vigorous shoots arise from unbombarded lateral meristems. Finally, an effective and versatile method for transgenic plant production was established by combining micro-wound treatment on a SAM by bombardment and LP-free SAM culture to suppress the growth of vigorous lateral shoots after wounding.
Mitsuru Sato; Takashi Kawabe; Munetaka Hosokawa; Fumi Tatsuzawa; Motoaki Doi
Plant cell reportsSPRINGER30(5)929 - 390721-77142011/05[Refereed]
The variegated Saintpaulia cultivar Thamires (Saintpaulia sp.), which has pink petals with blue splotches, is generally maintained by leaf cuttings. In contrast, tissue culture-derived progeny of the cultivar showed not only a high percentage of mutants with solid-blue petals but also other solid-color variants, which have not been observed from leaf cuttings. Solid-color phenotypes were inherited stably by their progeny from tissue culture. Petals from each solid-color variant were analyzed by high-performance liquid chromatography and shown to contain different proportions of three main anthocyanin derivatives: malvidin, peonidin, and pelargonidin. Analysis of flavonoid 3', 5'-hydroxylase (F3'5'H) sequences showed no differences in the coding region among the variants and variegated individuals. However, a transposon belonging to the hAT superfamily was found in the promoter region of variegated individuals, and the presence of transposon-related insertions or deletions correlated with the observed flower-color phenotypes. Solid-blue flower mutants contained 8-base pair (bp) insertions (transposon excision footprints), while solid-pink mutants had 58- to 70-bp insertions, and purple- and deep-purple mutants had 21- and 24-bp deletions, respectively. Real-time reverse transcription polymerase chain reaction (RT-PCR) analysis showed that F3'5'H expression levels correlated with insertions and deletions (indels) caused by hAT excision, resulting in flower-color differences. Our results showed that tissue culture of Saintpaulia 'Thamires' elicits transposon excision, which in turn alters F3'5'H expression levels and flower colors.
Song-Ji An; Devendra Pandeya; Soung-Woo Park; Jinjie Li; Jin-Kyung Kwon; Sota Koeda; Munetaka Hosokawa; Nam-Chon Paek; Doil Choi; Byoung-Cheorl Kang
TAG. Theoretical and applied genetics. Theoretische und angewandte GenetikSPRINGER122(3)459 - 700040-57522011/02[Refereed]
A temperature-sensitive mutant of Capsicum chinense, sy-2, shows a normal developmental phenotype when grown above 24°C. However, when grown at 20°C, sy-2 exhibits developmental defects, such as chlorophyll deficiency and shrunken leaves. To understand the underlying mechanism of this temperature-dependent response, phenotypic characterization and genetic analysis were performed. The results revealed abnormal chloroplast structures and cell collapse in leaves of the sy-2 plants grown at 20°C. Moreover, an excessive accumulation of reactive oxygen species (ROS) resulting in cell death was detected in the chlorophyll-deficient sectors of the leaves. However, the expression profile of the ROS scavenging genes did not alter in sy-2 plants grown at 20°C. A further analysis of fatty acid content in the leaves showed the impaired pathway of linoleic acid (18:2) to linolenic acid (18:3). Additionally, the Cafad7 gene was downregulated in sy-2 plants. This change may lead to dramatic physiological disorder and alteration of leaf morphology in sy-2 plants by losing low-temperature tolerance. Genetic analysis of an F(2) population from a cross between C. chinense 'sy-2' and wild-type C. chinense 'No. 3341' showed that the sy-2 phenotype is controlled by a single recessive gene. Molecular mapping revealed that the sy-2 gene is located at a genomic region of the pepper linkage group 1, corresponding to the 300kb region of the Ch1_scaffold 00106 in tomato chromosome 1. Candidate genes in this region will reveal the identity of sy-2 and the underlying mechanism of the temperature-dependent plant response.
Mitsuru Sato; Munetaka Hosokawa; Motoaki Doi
PloS onePUBLIC LIBRARY SCIENCE6(8)e23541 1932-62032011[Refereed]
BACKGROUND: The origin of somaclonal variation has not been questioned previously, i.e., "pre-existing mutations" in explants and "newly induced mutations" arising from the tissue culture process have not been distinguished. This is primarily because there has been no reliable molecular method for estimating or quantifying variation. METHODOLOGY/PRINCIPAL FINDINGS: We adopted a petal-variegated cultivar of Saintpaulia 'Thamires' (Saintpaulia sp.) as the model plant. Based on the difference between the pre- and post-transposon excision sequence of the promoter region of flavonoid 3', 5'-hydoroxylase (F3'5'H), we estimated mutated (transposon-excised) cell percentages using a quantitative real-time PCR. Mutated cell percentages in leaf laminae used as explants was 4.6 and 2.4% in highly or low variegation flower plants, respectively, although the occurrences of blue color mutants in their regenerants were more than 40%. Preexisting mutated cell percentages in cultured explants were considerably lower than the mutated plant percentage among total regenerants via tissue culture. CONCLUSIONS/SIGNIFICANCE: The estimation of mutated cell percentages became possible using the quantitative real-time PCR. The origins of mutations were successfully distinguished; it was confirmed that somaclonal variations are mainly caused by newly generated mutations arising from tissue culture process.
Yoshiyuki Tanaka; Munetaka Hosokawa; Tetsuya Miwa; Tatsuo Watanabe; Susumu Yazawa
Journal of agricultural and food chemistryAMER CHEMICAL SOC58(22)11762 - 70021-85612010/11[Refereed]
Capsinoids are a group of nonpungent capsaicinoid analogues produced in Capsicum fruits. They have similar bioactivities to capsaicinoids such as suppression of fat accumulation and antioxidant activity. They are more palatable ingredients in dietary supplements than capsaicinoids because of their low pungency. Previous studies on nonpungent Capsicum annuum cultivars showed that capsinoid biosynthesis is caused by loss-of-function putative aminotransferase (p-amt) alleles. This study showed that three mildly pungent cultivars of Capsicum chinense (Zavory Hot, Aji Dulce strain 2, and Belize Sweet) contain high levels of capsinoid. It was shown that these cultivars have novel p-amt alleles, which contain mutations that differ from those of C. annuum. Sequence analysis of p-amt in Belize Sweet revealed that a 5 bp insertion (TGGGC) results in a frameshift mutation. A transposable element (Tcc) was found in the p-amt of Zavory Hot and Aji Dulce strain 2. Tcc has features similar to those of the hAT transposon family. This was inserted in the fifth intron of Zavory Hot and in third intron of Aji Dulce strain 2. The p-amt alleles harboring Tcc cannot produce an active p-AMT. These mildly pungent cultivars will provide a new natural source of capsinoids.
Yoshiyuki Tanaka; Munetaka Hosokawa; Tetsuya Miwa; Tatsuo Watanabe; Susumu Yazawa
Journal of agricultural and food chemistryAMER CHEMICAL SOC58(3)1761 - 70021-85612010/02[Refereed]
Capsinoids make up a group of nonpungent capsaicinoid analogues produced in Capsicum fruits. They have bioactivities similar to those of capsaicinoids such as suppression of fat accumulation and antioxidant activity. Because of their low pungency, they are more palatable ingredients in dietary supplements than capsaicinoids. We recently reported that capsinoid biosynthesis is caused by nonsense mutation in a putative aminotransferase gene (p-AMT) in a nonpungent cultivar CH-19 Sweet. Here we report on the screening of nonpungent germplasm that revealed a nonpungent cultivar Himo, which contains high levels of capsinoids. We have shown that Himo has a recessive allele of p-amt, which contains a mutation different from that of CH-19 Sweet. Sequence analysis of p-amt in Himo revealed that a single-nucleotide substitution results in one amino acid substitution from cysteine to arginine in the pyridoxal 5-phosphate binding domain. Genetic analysis using a cleaved amplified polymorphic sequence marker confirmed that the p-AMT genotype was precisely cosegregated with capsinoid biosynthesis and nonpungency. Himo will provide a new natural source of capsinoids.
Yoshikuni Kitamura; Munetaka Hosokawa; Tatsuya Uemachi; Susumu Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV124(1)134 - 1380304-42382010/02[Refereed]
Hydrangea (Hydrangea spp.) has two types of florets in an inflorescence. One has decoratively developed sepals and is termed as the decorative floret. The other has plain sepals and is termed as the non-decorative floret. Hydrangea is classified into two types according to its inflorescence: globular (hortensia) and lacecap. In hortensia, the surface of inflorescence is covered with decorative florets. In lacecap, decorative florets are situated around the periphery of the inflorescence. Japanese hydrangea phyllody (JHP) phytoplasma infection often leads to an increase in the number of decorative florets. JHP phytoplasma was inoculated into ten hortensia and five lacecap cultivars by grafting. The ratios of decorative florets to total florets were compared between the JHP phytoplasma-infected and noninfected plants. The infected plants showed lower decorative floret ratios in six hortensia cultivars and higher decorative floret rations in four lacecap cultivars. The composition of inflorescence was investigated using infected and non-infected plants of 'Midoribanaajisai' (hortensia cultivar) and 'Libelle' (lacecap cultivar). In 'Libelle', the lacecap type, an alteration in the lateral non-decorative floret to the decorative floret was observed and considered to be the main cause of increase in the decorative floret ratio. (C) 2009 Elsevier B.V. All rights reserved.
伊藤弘顕; 西川久仁子; 粟野達也; 細川宗孝; 矢澤進
園芸学研究9(1)19-23 (J-STAGE) 2010[Refereed]
Yoshikuni Kitamura; Munetaka Hosokawa; Tatsuya Uemachi; Susumu Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV122(4)603 - 6090304-42382009/11[Refereed]
The mechanism underlying the green colouration of floral organs in phytoplasma-infected hydrangeas was studied. Elucidation of morphological changes of floral organs induced by phytoplasma infection and a comparison of the expression level of genes related to floral organ morphogenesis between phytoplasma-infected and non-infected hydrangea plants was conducted. Sixteen hydrangea (Hydrangea spp.) cultivars were used to study structural changes in floral organs induced by Japanese Hydrangea Phyllody (JHP) phytoplasma infection. The homeotic conversion of sepals to leaves was observed in different number of florets between cultivars, and was observed in 75.8-100% of the decorative florets and 2.7-67.3% of the non-decorative florets of JHP phytoplasma-infected inflorescences. The homeotic conversion of carpels to leaves was observed in 1.2-17.9% of the decorative florets and 0.1-18.9% of the non-decorative florets of infected inflorescences. The expression levels of hydrangea orthologues of class A, B and C genes were suppressed in the infected inflorescences during the developmental stage in which no morphological differences in the internal structure of floral organs have been observed. The contribution of this suppression to homeotic conversion of sepals and carpels to leaves was suggested. (C) 2009 Elsevier B.V. All rights reserved.
Sota Koeda; Munetaka Hosokawa; Byoung-Cheorl Kang; Susumu Yazawa
Journal of plant researchSPRINGER TOKYO122(6)623 - 310918-94402009/11[Refereed]
When a pepper cultivar (Capsicum chinense cv. Seychelles-2, Sy-2) native to the Seychelles was grown in Japan, all seedlings showed seasonal developmental abnormalities such as development of abnormally shaped leaves. Other pepper cultivars grew well in all seasons while the growth of cv. Sy-2 was stunted. In this study, we first examined the effects of various changes in temperature and photoperiod on the cv. Sy-2 phenotype. The results showed that temperatures lower than 24 degrees C led to the formation of abnormal leaves. Second, morphological and anatomical analyses of cotyledons and true leaves developed at 28 and 20 degrees C were conducted. The narrower and thicker cotyledons developed at 20 degrees C had fewer palisade cells in the leaf-length direction, and more cells in the leaf-thickness direction. True leaves developed at 20 degrees C were irregularly shaped, thicker and had smaller leaf area. In addition, true leaves developed at 20 degrees C had fewer palisade cells in the leaf-length and leaf-width directions and had more cells in the leaf-thickness direction. Furthermore, abnormal periclinal cell divisions in the mesophyll and/or epidermal cell layers were observed during leaf blade development at 20 degrees C. These results suggest that the observed changes in cell proliferation and abnormal periclinal cell divisions were related, at least in part, to abnormal leaf development of cv. Sy-2 at temperatures below 24 degrees C.
細川宗孝; 大野翔; 北村嘉邦; 児島未沙; 中島明子; 星野敦; 立澤文見; 土井元章
園芸学研究 別冊8(2)528 1881-83072009/09[Refereed]
Hisanao Suzue; Munetaka Hosokawa; Susumu Yazawa
HORTSCIENCEAMER SOC HORTICULTURAL SCIENCE44(5)1501 - 15030018-53452009/08[Refereed]
Exposing the shoot apical meristem (SAM) is necessary in horticultural practices such as shoot tip culture, colchicine treatment, gene transfer, and so on. However, ordinary methods of exposing SAMs require extensive training and are labor-intensive. We developed equipment to expose SAMs quickly and easily by blowing compressed air on the shoot tips. Using this equipment allowed SAMs to be exposed more rapidly than the ordinary methods. Exposure time was shortened from 27.8 s to 7.4 s, 15.7 s to 7.9 s, and 59.7 s to 6.8 s in chrysanthemum (Chrysanthemum morifolium), dahlia (Dahlia spp.), and Lilium leichtlinii, respectively. To determine whether blowing compressed air caused injury to SAMs, we used shoots in which the SAMs had been already exposed and observed resin sections of SAMs that had been subjected to the additional compressed air treatment for several seconds. Results showed that blowing additional compressed air for as long as 9 s did not injure SAMs. Using this equipment, we can expose SAMs without using a stereomicroscope by blowing compressed air to the top of shoots for several seconds. Over 70% of SAMs could he exposed in a 7- to 13-s blowing treatment without injury. We could successfully use this equipment in shoot tip culture and leaf primordia-free SAM culture.
Hitomi Omori; Munetaka Hosokawa; Hayato Shiba; Naoki sh*tsukawa; Koji Murai; Susumu Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI78(3)350 - 3551882-33512009/07[Refereed]
Chrysanthemum stunt viroid (CSVd) is one of the problematic pathogens known to infect chrysanthemum (Dendranthema grandiflorum) plants and causes various symptoms, such as a reduction in plant height, which is a serious problem in cut flower production. No natural sources with resistance to CSVd have been reported. By quantifying the CSVd titer within the plant, we identified the cultivar 'Utage' as a plant in which the increase of the CSVd titer was slowest among 6 cultivars tested. 'Utage' self-pollinated, and 67 resulting seedlings were screened for CSVd resistance using reverse transcription polymerase chain reaction (RT-PCR), nested-PCR, micro-tissue (MT) direct RT-PCR, and real-time RT-PCR. Of these 67 seedlings, 9 plants lacked the obvious CSVd band detected by RT-PCR. Five months after grafting to CSVd-infected plants, the CSVd titers of 3 plants (C7, A30, and A27) were about 1/240, 1/41000, and 1/125000 compared to that of 'Utage', respectively. These 3 plants were comfirmed to have strong resistance to CSVd. In C7, local distribution of CSVd was observed in the youngest expanded leaf by micro-tissue direct RT-PCR and in situ hybridization. For A30 and A27, CSVd was hardly detected in the whole plant. These 3 plants will contribute to the elucidation of CSVd resistance mechanisms.
Yoshiyuki Tanaka; Munetaka Hosokawa; Keigo Otsu; Tatsuo Watanabe; Susumu Yazawa
Journal of agricultural and food chemistryAMER CHEMICAL SOC57(12)5407 - 120021-85612009/06[Refereed]
Capsiconinoid is a group of nonpungent capsaicinoid analogues produced in Capsicum fruits, which we recently identified. Capsiconinoids have agonist activity for transient receptor potential vanilloid type 1 (TRPV1), which is reported to be a receptor for capsaicin. It is, therefore, important to screen cultivars containing high levels of capsiconinoid for their use as a vegetable or dietary supplement. This study describes the quantitative analysis of capsiconinoid content in fruits of 35 Capsicum cultivars: 18 cultivars of C. annuum, 7 of C. baccatum, 5 of C. chinense, 4 of C. frutescens, and 1 of C. pubescens. Using high-performance liquid chromatography (HPLC), we found that 10 cultivars contained capsiconinoids. Capsiconinoid Baccatum (CCB) (C. baccatum var. praetermissum) showed the highest capsiconinoid content (3314 microg/g DW) and Charapita (C. chinense) had the second highest content. The other 8 cultivars had much lower capsiconinoid content than these two cultivars (<300 microg/g DW). Time-course analysis during fruit development clarified that capsiconinoid content in CCB fruits increased until 30 days after flowering (DAF) and then decreased rapidly until 40 DAF.
Yang Zhang; Takahiro Hayashi; Munetaka Hosokawa; Susumu Yazawa; Yuhua Li
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV121(2)213 - 2170304-42382009/06[Refereed]
The mechanism of sparkling metallic lustre formation in leaves of Begonia rex Putz. was investigated. On the metallic lustre and normal green areas of one leaf, we, observed two light reflection patterns, one a spotted pattern (SP) composed of white spots formed on the centre of epidermal cells and the other a polygonal pattern (PP) composed of white polygons formed around the epidermal cell edges. The SP was observed both in metallic lustre and normal green areas, and the intensity did not differ significantly, but the PP was much stronger in the metallic lustre area. Different reflection of the PP in metallic lustre and normal green areas indicated that metallic lustre formation was attributable to the PP. Light reflection of the PP was mainly derived from interior light reflection because the PP became very weak after air removal from leaf. The formation of the PP is associated with the interior structure of the leaf, such as chlorophyll density, cell arrangement and air space. The experimental results indicated that interior air space is an important factor in forming the polygonal pattern. (c) 2009 Elsevier B.V. All rights reserved.
Elly Kesumawati; Munetaka Hosokawa; Takushi Kimata; Tatsuya Uemachi; Susumu Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV121(2)199 - 2050304-42382009/06[Refereed]
To determine the effect of light intensity on flower greening, the Japanese hydrangea phyllody (JHP) phytoplasma-infected hydrangea cultivars 'Midori', 'Libelle', 'Rosea' and 'Madame E. Mouillere' plants were grown under different shade conditions. In the first-year experiment, the results indicate that the flowers of the JHP-phytoplasma-infected hydrangea become green under shaded conditions (70% and 49% sunlight intensities). On the other hand, under full sunlight intensity (100% sunlight intensity), the flowers of 'Midori','Rosea', and 'Libelle' plants were blue, pink or white. To calculate the percentage of flower greening, inflorescences of these plants were separated and divided into individual flowers, and classified into four types by green-area ratio, calculated using Adobe Photoshop. Under shading with one sheet of cheesecloth (70% sunlight intensity), the inflorescences of 'Midori', 'Libelle' and 'Madame E. Mouillere' plants were composed of more than 40% completely green flowers (0.8 <= green-area ratio), whereas those of 'Rosea' plant had 0% completely green flowers. Under shading with two sheets of cheesecloth (49% sunlight intensity), the inflorescences of 'Midori', 'Libelle' and 'Madame E. Mouillere' plants had more than 75% completely green flowers; 'Rosea' plants had 28%. In the second-year experiment, under full sunlight intensity, 'Midori' plants had four types of flower depending on their green-area ratio, namely, completely blue or pink, pink-green, greenish and completely green flowers. Under shading with two sheets of cheesecloth, 'Midori' plants had more than 90% completely green flowers. The JHP-phytoplasma could not be identified by PCR analysis in flowers with a green-area ratio = 0 (completely blue/pink/white flowers). On the other hand, in flowers with a green-area ratio > 0, the JHP-phytoplasma was detected by PCR analysis. Thus, we conclude that shading enhances flower greening in hydrangea by increasing the JHP-phytoplasma concentration in the flowers. (c) 2009 Elsevier B.V. All rights reserved.
Yoshikuni Kitamura; Munetaka Hosokawa; Chihiro Tanaka; Susumu Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI77(4)418 - 4251882-33512008/10[Refereed]
In some plant species, bacterial contamination frequently occurs in hi vitro culture. Also, in hydrangea (Hydrangea spp.), microbial contamination frequently occurs when their explants are cultured in vitro. We identified bacterial flora near hydrangea shoot apical meristems (SAMs) by analyzing their 16S ribosomal DNA (16S rDNA) sequences. Sequences of 16S rDNA fragments amplified from bacteria isolated from SAMs of 8 hydrangea cultivars were identical to those of 12 bacterial species. Because I to 9 bacterial species were detected only once in each cultivar, and we focused on bacteria that can colonize on Nutrient Broth medium, in this reserch we could not cover all bacterial species existing near SAMs. The appropriate chlorine concentration for sterilizing SAMs was decided using 'Miss Hepburn'. When shoot tips with stripped SAMs were dipped in chlorine solution with 0.0005%-0.5% available chlorine concentration for 30 min, no bacterial colony was observed in the treatment with 0.05% chlorine. From this result, direct exposure of the stripped meristem to chlorine solution is a suitable method for sterilizing hydrangea shoot tip culture explants, and the bacteria causing in vitro contamination in hydrangea shoot-tip culture are not endophytic butepiphytic. A successful method for in vitro culture of hydrangea was established using 8 hydrangea cultivars in Exp. 3 and in 7 cultivars, we could gain 5 to 19 viable plants from 20 cultured explants without bacterial contamination by sterilizing shoot tips for 30 min with a sterilization solution of 0.05% available chlorine concentration after removal of all but two pairs of leaf primordia. No viable explant was gained from 'Flambeau', so the chlorine concentration must be reconsidered to sterilize such cultivars that have chlorine-sensitive meristems. One hundred and thirty days after culture initiation, no contamination was observed and cultured explants grew sufficiently to be transplanted out of the bottle. Surface sterilization of the SAM can be applied for many plants in which bacterial contamination poses a big problem in shoot-tip culture.
Leaf Primordia-Free Shoot Apical Meristem Culture: A New Method for Production of Viroid-Free Plants
Munetaka Hosokawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI77(4)341 - 3491882-33512008/10[Refereed]
Viroids are the smallest and one of the most problematic pathogens known in horticulture. Despite not coding for proteins they replicate in nuclei or chloroplasts using host enzymes. From many pathological studies, it is obvious that viroids are completely different agents from viruses. For the past 40 years, elimination of viroids from infected plants has been the main theme in horticulture and several approaches have been established; however, these procedures remain complicated. Leaf primordia-free shoot apical meristem culture is a newly established method that can effectively eliminate viroids. Therefore, in this review, the method is summarized and the mechanism of viroid elimination is discussed.
Kuniko Nishikawa; Hiroaki Ito; Tatsuya Awano; Munetaka Hosokawa; Susumu Yazawa
Annals of botanyOXFORD UNIV PRESS102(1)31 - 70305-73642008/07[Refereed]
BACKGROUND AND AIMS: Helichrysum bracteatum is called an 'eternal flower' and has large, coloured, scarious bracts. These maintain their aesthetic value without wilting or discoloration for many years. There have been no research studies of cell death or cell morphology of the scarious bract, and hence the aim of this work was to elucidate these characteristics for the bract of H. bracteatum. METHODS: DAPI (4'6-diamidino-2-phenylindol dihydrochloride) staining and fluorescence microscopy were used for observation of cell nuclei. Light microscopy (LM), transmission electron microscopy (TEM) and polarized light microscopy were used for observation of cells, including cell wall morphology. KEY RESULTS: Cell death occurred at the bract tip during the early stage of flower development. The cell wall was the most prominent characteristic of H. bracteatum bract cells. Characteristic thickened secondary cell walls on the inside of the primary cell walls were observed in both epidermal and inner cells. In addition, the walls of all cells exhibited birefringence. Characteristic thickened secondary cell walls have orientated cellulose microfibrils as well as general secondary cell walls of the tracheary elements. For comparison, these characters were not observed in the petal and bract tissues of Chrysanthemum morifolium. CONCLUSIONS: Bracts at anthesis are composed of dead cells. Helichrysum bracteatum bracts have characteristic thickened secondary cell walls that have not been observed in the parenchyma of any other flowers or leaves. The cells of the H. bracteatum bract differ from other tissues with secondary cell walls, suggesting that they may be a new cell type.
Y. Zhang; T. Hayashi; M. Inoue; Y. Oyama; M. Hosokawa; S. Yazawa
PROCEEDINGS OF THE INTERNATIONAL SYMPOSIUM ON ORNAMENTALS, NOW!INTERNATIONAL SOCIETY HORTICULTURAL SCIENCE766(766)469 - 4750567-75722008[Refereed]
The textures of flowers are important ornamental characteristics. Reflected light from the surface of petals, including colorful bracts and leaves, is a key factor determining the texture of flowers. The results of experiments in which we examined various species of flower petals indicated that there are two types of reflected light: one is superficial reflected light, which is determined by the shape of the epidermal cells. More light is reflected by flat epidermal cells than by papillate ones. In particular, when the angle of incident light was changed, the belt-shaped reflected light was only observed through the microscope from the side of the papillate epidermal cells. The other type of reflected light is scattered reflected light, which is determined by the petal structure, and the higher the pigment content in a petal, the more light is absorbed. The intensity and origin of scattered reflected light depends on the volume and location of air spaces in the petal that are distributed among the epidermal, palisade, and spongy cells. There are three typical flower textures: velvet luster (e.g., Viola tricolor), which occurs due to superficial reflected light, and metallic luster (e.g., Begonia rex) and diamond dust (e.g., Saintpaulia ionantha), which are caused by scattered reflected light.
Akiko Nakashima; Munetaka Hosokawa; Shigeichi Maeda; Susumu Yazawa
Journal of General Plant Pathology73(3)225 - 2271345-26302007/06[Refereed]
For the first time, Chrysanthemum stunt viroid (CSVd) was detected in commercial dahlia bulbs in Japan. CSVd was found in 77.2% of the tested plants (Dahlia spp.). In nucleotide sequence analysis, a CSVd variant was detected consisting of 354 nucleotides, which differed slightly from previously reported CSVd variants. © 2007 The Phytopathological Society of Japan and Springer.
Munetaka Hosokawa; Hayato Shiba; Takashi Kawabe; Akiko Nakashima; Susumu Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI76(1)60 - 651882-33512007/01[Refereed]
The economically important Chrysanthemum stunt viroid (CSVd) and Chrysanthemum chlorotic mottle viroid (CChMVd) were detected from infected chrysanthemum (Dendranthema grandiflorum) plants by a multiplex reverse transcription-polymerase chain reaction (RT-PCR) assay developed in this investigation. The antisense hexamer AAAGGA (5'-3') was designed. The antisense hexamer 5'AAAGGA 3' annealing at 5'TCCTTT3' is located at nucleotide positions 186-191 and 245-250 of CSVd (gb: AB006737), and 231-236 of CChMVd in their sequences. Using CSVd- and CChMVd-cDNA templates which were transcribed simultaneously by the hexamer, the following multiplex PCR could detect both viroids from doubly-infected plants without nonspecific amplification. When the hexamer was used for the RT reaction, the sensitivity of detection of CSVd or CChMVd by multiplex RT-PCR was similar to that of standard RT-PCR when each viroid was detected separately. Furthermore, multiplex RT-PCR successfully detected both CSVd and CChMVd in direct templates obtained by inserting a syringe needle into the stem, leaf, and shoot tips of infected chrysanthemum plants. The direct multiplex RT-PCR method developed in this study may reduce the cost, time, and labor required for the production of viroid-free chrysanthemum plants.
Soo Jung Yang; Yoshikuni Kitamura; Munetaka Hosokawa; Susumu Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI75(6)476 - 4801882-33512006/11[Refereed]
Hypersensitivity is induced in many species of Acanthaceae and Gesneriaceae by local mechanical wounding. Once a plant of these species sustains a wounded leaf, it exhibits a temporary hypersensitive state and severe injury easily occurrs even in unwounded leaves. Locally wounded African violet (Saintpaulia sp.) plants sometimes show severe injury even at a noninjurious temperature for this species. This study was aimed to clarify changes in the temperature sensitivity of hypersensitivity-induced African violet plants by local wounding. The basal leaves of African violet 'Ritali' plants were cut as a local wounding treatment. The locally wounded plants were transferred to growth chambers at various temperatures and we evaluated the degree of leaf injury using an analysis software. In locally wounded plants, temperature sensitivity increased and leaf injury was observed even at 11 degrees C, which is a noninjurious temperature for unwounded African violet. Such a wound-induced sensitivity to low temperature treatment (7 degrees C for 1h) quantitatively increased with local wounding severity. For the investigation of time-course changes in the hypersensitive state (wounding memory) of locally wounded plants, the plants were transferred to low temperature conditions at various intervals after the wounding treatment. The wounding memory was short-lived, that is, it was generally observed immediately after local wounding and then almost disappeared 30 min after. In conclusion, the low temperature sensitivity of African violet plants was enhanced by prewounding treatment.
E Kesumawati; T Kimata; T Uemachi; M Hosokawa; S Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV108(1)74 - 780304-42382006/03[Refereed]
The interaction between phytoplasma concentration and green-flowering stability was studied in hydrangea cultivars. Three green and 18 nongreen cultivars were subjected to polymerase chain reaction (PCR) analysis to determine Japanese hydrangea phyllody (JHP) phytoplasma infection. The results showed that JHP-phytoplasma was detected only in 'Midori' plants, which have green sepal, 'Midori' plants were propagated, and from 29 rooted cutting plants, they were grouped into three types on the basis of sepal color. that is. green (75,9%). blue-green (13.8%) and blue (10.3%) sepals. To clarify the variability in the sepal color of 'Midori' plants. JHP-phyloplasma concentration in the sepals and leaves of green-, blue-green- and blue-flowering plants was determined by PCR analysis, The semiquantitative PCR comparisons of 370 bp DNA fragments showed that the JHP-phytoplasma concentrations in green sepals were 16 times higher than that in blue-green sepals, JHP-phytoplasma could not be identified by PCR analysis in blue sepals and leaves. These results showed that JHP-phytoplasma concentration correlated with green sepal stability in 'Midori' plants. A histological observation of sepals showed that epidermal cells of blue and blue-green sepals had a dome shape. Otherwise, green sepals were leaflike with flat epidermal cells, and palisade parenchyma cells with numerous, chloroplasts. (c) 2006 Elsevier B.V. All rights reserved.
M Hosokawa; Y Matsush*ta; H Uchida; S Yazawa
JOURNAL OF VIROLOGICAL METHODSELSEVIER SCIENCE BV131(1)28 - 330166-09342006/01[Refereed]
A direct reverse transcription-polymerase chain reaction (RT-PCR) method for detecting the chrysanthemum stunt viroid (CSVd) and chrysanthemum chlorotic mottle viroid (CChMVd) to screen for a viroid-free chrysanthemum plant at a small plant size was established and named microtissue direct RT-PCR. A razor or syringe needle was used for RNA template preparations. Under a stereoscopic microscope, a razor or syringe needle was used to pierce, a tissue sample to a depth of 0.1-0.2 mm, and the sample was directly transferred to the RT mixtures. Methods using razors or needles for the preparation of templates could detect CSVd and CChMVd with a high sensitivity. The most sensitive method used a razor or syringe needle to acquire template from the shoot tips. Using the microtissue direct RT-PCR method, both viroids could be detected from the high- and low-viroid-concentration plants. The microtissue direct RT-PCR method was more sensitive than a conventional template preparation method. Using the microtissue direct RT-PCR method established in this study, the laborious subculture step could be omitted because detecting viroids and screening for viroid-free plants even at a small plant size before the subculture could be possible. In addition, the microtissue direct RT-PCR method could also be a powerful tool for clarifing the viroid distribution among microtissues, such as shoot apical meristems. (c) 2005 Elsevier B.V. All rights reserved.
M Hosokawa; Y Matsush*ta; K Ohishi; S Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI74(5)386 - 3911882-33512005/09[Refereed]
The disease symptoms of chrysanthemum chlorotic mottle viroid (CChMVd) that had not been observed in chrysanthemum (Dendranthema grandiflorum) plants grown in Japan were found recently. In our first experimental test to examine whether the disease symptoms, such as chlorosis of newly expanded leaves in Japanese chrysanthemum, confirmed that the disease was induced by CChMVd. The viroid was detected in cultivars with the disease symptoms by polymerase chain reaction (PCR). The nucleotide sequence of CChMVd extracted from an infected 'Piato' plant, almost completely agreed with that of the non-symptomatic CChMVd strain that was reported previously. In the second experiment from cultivars randomly collected from various areas in Japan, namely, f*ckuoka, Hiroshima, Osaka, Shiga, Kyoto and Aichi prefectures, CChMVd was detected in many of the collected cultivars. Moreover, CChMVd was detected in several samples at low concentrations from cut chrysanthemum flowers sold in Kyoto prefecture shipped from various areas in Japan and the Netherlands. Hence, we concluded that CChMVd had already spread in Japan. In the third experiment to develop a CChMVd- free plant through a newly established method, that is, leaf primordia-free shoot apical meristem (LP-free SAM) culture was utilized to eliminate CChMVd from infected cultivars. These excised LP- free SAMs derived from CChMVd -infected 'Piato' and 'Sttetsuman' plants were attached to the root tip of in vitro-sown cabbage (Brassica oleracea) 'Harunami' and regenerated. One out of 29 regenerated 'Piato' plants and two out of 6 regenerated 'Sttetsuman' plants were ascertained to be CChMVd -free by nested PCR.
Munetaka Hosokawa; Emi Ueda; Kazushi Ohishi; Ayaka Otake; Susumu Yazawa
PlantaSPRINGER220(1)64 - 700032-09352004/11[Refereed]
Chrysanthemum ( Dendranthema grandiflorum Kitam.) is one of the qualitative short-day flowering plants. Therefore, the flowering of chrysanthemum can usually be controlled by photoperiod. However, it was noted that 'Piato' plants infected by the chrysanthemum stunt viroid (CSVd) flowered autonomously even under long-day conditions. In this study, CSVd-free and CSVd-infected plants were prepared by culturing different-sized dissected shoot apical meristems (SAMs) of 'Piato'. Using these CSVd-free and CSVd-infected plants, we clarified the relationship between CSVd infection and the autonomous flowering of 'Piato'. Under natural short-day conditions, the flowering of plants regenerated from SAMs containing leaf primordia (LPs) was 1 month earlier than plants regenerated from LP-free SAMs. CSVd was detected from these early flowering plants by reverse transcription-polymerase chain reaction. On the other hand, CSVd was not detected in plants regenerated from LP-free SAMs. CSVd-infected and CSVd-free plants were grown under long-day conditions simulated by night-break lighting at 22:00 p.m. to 2:00 a.m. All CSVd-infected plants flowered autonomously even under long-day conditions; on the other hand, CSVd-free chrysanthemum plants maintained their vegetative growth. When the CSVd-free plants were inoculated with CSVd by grafting them to CSVd-infected rootstocks, they flowered autonomously even under night-break lighting. In this study, the results suggest that CSVd may control the qualitative development process, flowering, i.e. CSVd can induce the autonomous flowering of chrysanthemum.
M Hosokawa; A Otake; K Ohishi; E Ueda; T Hayashi; S Yazawa
PLANT CELL REPORTSSPRINGER-VERLAG22(11)859 - 8630721-77142004/06[Refereed]
In this research we eliminated chrysanthemum stunt viroid (CSVd) from a highly infected chrysanthemum cultivar using a newly established method. 'Piato' is one of the most difficult cultivars in which to obtain CSVd-free plants by conventional methods. Leaf primordium-free shoot apical meristems (LP-free SAMs) of 'Piato' plants were dissected and attached to CSVd-free chrysanthemum or cabbage root tips. As shown by nested-PCR, CSVd was not detected in some shoots regenerated on both types of root tip. The production rates of CSVd-free plants using chrysanthemum and cabbage root tips were 14% and 3%, respectively. Regeneration of plants from LP-free SAMs of chrysanthemum plants by attaching these SAMs to root tips is an efficient method of generating CSVd-free chrysanthemum plants.
M Hosokawa; A Otake; Y Sugawara; T Hayashi; S Yazawa
PLANT CELL REPORTSSPRINGER-VERLAG22(7)443 - 4480721-77142004/02[Refereed]
A new method to regenerate plants from leaf primordia-free shoot apical meristem domes (LP-free SAMs) was developed by establishing the meristem dome on the cut surface of root tips. Ten days after culture, the viable rate of LP-free SAMs of chrysanthemum 'Piato' attached to chrysanthemum root tips was >40%. Shoot regeneration was not observed from LP-free SAMs without the root tips. When LP-free SAMs of chrysanthemum were transferred to root tips of either petunia, cabbage, or carnation, the highest shoot regeneration rate was observed with cabbage root tips. Microscopic observation documented that the LP-free SAM temporarily adhered to the cut surface of the root tip of cabbage.
SJ Yang; M Hosokawa; T Hayashi; S Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI72(4)286 - 2911882-33512003/07[Refereed]
To evaluate the effect of wounding on the incidence of leaf browning to a sudden temperature drop, Ruellia macrantha, cuttings of a highly wound-sensitive Acanthaceae species, were briefly immersed in water (3- 9 degreesC lower than the leaf temperature for 3 sec). Leaf browning occurred in wounded plants but not in intact ones. The incidence and the severity of leaf browning increased with decreased temperatures, but even the immersion in 25 degreesC water induced leaf browning in injured plants (25 degreesC). These responses indicate that the water stimulates leaf browning in a wounded plant, whereas the reduced temperature intensifies the responsiveness. This wound-induced increase in sensitivity to water stimulus is referred to as hypersensitivity. Injuring the leaf, petiole, stem or root by cutting or crushing induced hypersensitivity only in young expanded leaves above the wound site. The hypersensitivity reached maximum 2 or 3 min after wounding and disappeared after 30 min. In addition, cutting the shoot a short distance from the apex resulted in severe leaf browning, indicating that the distance from young expanded leaves to the site of wounding affects the level of hypersensitivity. Our data indicate that a wound signal is rapidly translocated to the unwounded leaves to play a role in inducing hypersensitive responses to mechanical stimuli, such as water stimulus and sudden temperature drop.
EU Ahmed; T Hayashi; Y Zhu; M Hosokawa; S Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV96(1-4)187 - 1940304-42382002/12[Refereed]
Explants from various parts of Caladium bicolor Ait. plants were cultured on MS medium containing 1 ppm naphthalene acetic acid (NAA) and 1 ppm 6-benzyladenine (BA). Variations in the shape and color pattern were observed in the regenerated plants. In plants regenerated from shoot tip explants including the two youngest leaves, young leaves (third and fourth leaves from the top), unexpanded leaves (fifth or sixth leaves), expanded leaves and petiole, the frequency of the occurrence of variants was 14, 24, 41, 53 and 70%, respectively. Plants regenerated from young leaves of lateral buds or root tips also produced only a few variants (12-14%). Thus, explants from younger tissue having meristematic tissue produced fewer variations than explants from older tissue. The mode of variation (color pattern and shape of leaves) also varied with the age of the explant. We conclude that young tissue should be used to propagate Caladium plants by tissue culture. (C) 2002 Elsevier Science B.V. All rights reserved.
S Ahmed; E Nawata; M Hosokawa; Y Domae; T Sakuratani
PLANT SCIENCEELSEVIER SCI IRELAND LTD163(1)117 - 1230168-94522002/07[Refereed]
Photosynthetic rate (P), leaf water potential (psi(L)) and chlorophyll fluorescence were studied in mungbean plants subjected to waterlogging in a plastic greenhouse, and leaf samples were collected for the analysis of enzymes involved in anti-oxidant photoprotection. Waterlogging caused a fast decline in the photosynthetic rate and water use efficiency without changes of transpiration rate and stomatal conductance during the early waterlogging period, which indicates that reduction of photosynthetic rate might be due to a mechanism independent from stomatal closure. Early reduction of Fv/Fm value of chlorophyll fluorescence by waterlogging indicates that the early reduction of photosynthetic rate is correlated with photo inhibition of photosystem II (PSII). Stomatal conductance and water use efficiency was decreased progressively during prolonged waterlogging while psi(L) was unchanged, indicating stomatal closure without water stress. The activities of active oxygen processing enzymes, such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) decreased during prolonged waterlogging indicating no evidence of oxidative stress and/or damage in leaves of mungbean by waterlogging. Our results suggest that oxidative damage is not directly involved in damage to the photosynthetic machinery during waterlogging in mungbean plants. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.
SJ Yang; M Hosokawa; T Hayashi; S Yazawa
JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL SCIENCEJAPAN SOC HORTICULTURAL SCI71(4)535 - 5371882-33512002/07[Refereed]
Leaf browning was induced at sites distant from the wounds in detached leaves in 6 species of Acanthaceae and 13 species of Gesneriaceae plants. This injury was observed in unwounded young leaves after their detachment from the mother plant, and sometimes in unwounded leaves on the wounded plant, e.g., when stem cuttings were made. The injury is restricted to one layer of palisade cells; the response is rapid. Chlorophyll fluorescence intensity decreased irreversibly in an injured tissue within seconds at sites distant from the wound.
SJ Yang; M Hosokawa; Y Mizuta; JG Yun; J Mano; S Yazawa
SCIENTIA HORTICULTURAEELSEVIER SCIENCE BV88(1)59 - 690304-42382001/03[Refereed]
The relationship between the susceptibility to rapid temperature drop induced leaf injury, i.e. "leaf spot", and the antioxidant capacities of several Saintpaulia (African violet) cultivars was examined. Cultivars 'Ritali' and 'Tamiko', that are more susceptible to leaf spot caused by a rapid drop in leaf temperature from 30 to 15 degreesC than are cultivars 'Maui' and 'New Jersey'. The susceptible cultivars were also more susceptible to oxidative stress caused by H2O2 and active chlorine than the tolerant cultivars. Reduction of available chlorine, considered to be accelerated by antioxidants of leaf tissue, was rapid in 'Maui' and 'New Jersey'. Activity of the antioxidant enzymes superoxide dismutase and catalase were higher in the leaves of 'Maui' and 'New Jersey' than in the susceptible cultivars. These findings show a correlation between leaf spot injury and antioxidant capacity in Saintpaulia leaves. (C) 2001 Elsevier Science B.V. All rights reserved.
細川 宗孝
Plant Tissue Culture and Biotechnology,4/1,35435 - 411998[Refereed]
