Hiroshi Ishizaka

Publications (3)6.14 Total impact

• Article: Molecular characterization of an anthocyanin-related glutathione S-transferase gene in cyclamen.

  Satoshi Kitamura, Yusuke Akita, Hiroshi Ishizaka, Issay Narumi, Atsushi Tanaka
  [show abstract] [hide abstract]
  ABSTRACT: Anthocyanins are a subclass of flavonoids and are a major contributor to flower colors ranging from red to blue and purple. Previous studies in model and ornamental plants indicate a member of the glutathione S-transferase (GST) gene family is involved in vacuolar accumulation of anthocyanins. In order to identify the anthocyanin-related GST in cyclamen, degenerate PCR was performed using total RNA from immature young petals. Four candidates of GSTs (CkmGST1 to CkmGST4) were isolated. Phylogenetic analysis indicated that CkmGST3 was closely related to PhAN9, an anthocyanin-related GST of petunia, and this clade was clustered with other known anthocyanin-related GSTs. Expression analysis at different developmental stages of petals revealed that CkmGST3 was strongly expressed in paler pigmented petals than in fully pigmented petals, in contrast to the constitutive expression of the other three candidates during petal development. This expression pattern of CkmGST3 was correlated with those of other anthocyanin biosynthetic genes such as CkmF3'5'H and CkmDFR2. Molecular complementation of Arabidopsis tt19, a knockout mutant of an anthocyanin-related GST gene, demonstrated that CkmGST3 could complement the anthocyanin-less phenotype of tt19. Transgenic plants that expressed the other three CkmGSTs did not show anthocyanin accumulation. These results indicate CkmGST3 functions in anthocyanin accumulation in cyclamen.
  Journal of plant physiology 04/2012; 169(6):636-42. · 2.50 Impact Factor
• Article: Isolation and characterization of the fragrant cyclamen O-methyltransferase involved in flower coloration.

  Yusuke Akita, Satoshi Kitamura, Yoshihiro Hase, Issay Narumi, Hiroshi Ishizaka, Emiko Kondo, Naoko Kameari, Masayoshi Nakayama, Natsu Tanikawa, Yasumasa Morita, Atsushi Tanaka
  [show abstract] [hide abstract]
  ABSTRACT: Anthocyanin O-methyltransferase (OMT) is one of the key enzymes for anthocyanin modification and flower pigmentation. We previously bred a novel red-purple-flowered fragrant cyclamen (KMrp) from the purple-flowered fragrant cyclamen 'Kaori-no-mai' (KM) by ion-beam irradiation. Since the major anthocyanins in KMrp and KM petals were delphinidin 3,5-diglucoside and malvidin 3,5-diglucoside, respectively, inactivation of a methylation step in the anthocyanin biosynthetic pathway was indicated in KMrp. We isolated and compared OMT genes expressed in KM and KMrp petals. RT-PCR analysis revealed that CkmOMT2 was expressed in the petals of KM but not in KMrp. Three additional CkmOMTs with identical sequences were expressed in petals of both KM and KMrp. Genomic PCR analysis revealed that CkmOMT2 was not amplified from the KMrp genome, indicating that ion-beam irradiation caused a loss of the entire CkmOMT2 region in KMrp. In vitro enzyme assay demonstrated that CkmOMT2 catalyzes the 3' or 3',5' O-methylation of the B-ring of anthocyanin substrates. These results suggest that CkmOMT2 is functional for anthocyanin methylation, and defective expression of CkmOMT2 is responsible for changes in anthocyanin composition and flower coloration in KMrp.
  Planta 07/2011; 234(6):1127-36. · 3.00 Impact Factor
• Article: Comparative analysis of floral pigmentation between wild-type and white-flowered varieties of Cyclamen graecum

  Yusuke Akita, Hiroshi Ishizaka, Masayoshi Nakayama, Akihiko Shimada, Satoshi Kitamura, Yoshihiro Hase, Issay Narumi, Atsushi Tanaka
  [show abstract] [hide abstract]
  ABSTRACT: The flower colour of Cyclamen graecum gra6 (wild-type) is pink-purple in the main part of the petal, referred to as the ‘slip’, and deep purple at the petal base, referred to as the ‘eye’. On the other hand, flowers of C. graecum gra50 (a white-flowered variant) exhibit a white colour in both the ‘slip’ and ‘eye’ regions. In this study, the relationship between floral pigmentation and the expression of several anthocyanin biosynthesis genes was investigated in C. graecum gra6 and gra50.The pigments in the ‘slip’ and ‘eye’ regions consist mainly of malvidin 3,5-diglucoside in gra6, suggesting that the difference between the colour of the ‘slip’ and ‘eye’ regions is related to the amount of anthocyanin present.White-flowered C. graecum gra50 possessed lower amounts of anthocyanins, but higher amounts of flavonols compared to gra6, suggesting a change in metabolism caused by a disruption of anthocyanin biosynthesis. Gene expression analysis demonstrated that expression of the dihydroflavonol 4-reductase gene 2 (CgraDFR2) was lower in gra50 compared with gra6, whereas expression of the three other key genes (dihydroflavonol 4-reductase gene 1, flavonoid 3’,5’-hydroxylase, and anthocyanidin synthase) did not differ greatly. These results suggest that the white-flowered variant (gra50) may result from a defect in expression of the CgraDFR2 gene.
  Journal of Horticultural Science and Biotechnology 05/2010; 85(5):437-43. · 0.64 Impact Factor