Kazuyuki Abe

Molecular Characterization of FLOWERING LOCUS T-Like Genes of Apple (Malus × domestica Borkh.)

The two FLOWERING LOCUS T (FT)-like genes of apple (Malus x domestica Borkh.), MdFT1 and MdFT2, have been isolated and characterized. MdFT1 and MdFT2 were mapped, respectively, on distinct linkage groups (LGs) with partial homoeology, LG 12 and LG 4. The expression pattern of MdFT1 and MdFT2 differed in that MdFT1 was expressed mainly in apical buds of fruit-bearing shoots in the adult phase, with little expression in the juvenile tissues, whereas MdFT2 was expressed mainly in reproductive organs, including flower buds and young fruit. On the other hand, both genes had the potential to induce early flowering since transgenic Arabidopsis, which ectopically expressed MdFT1 or MdFT2, flowered earlier than wild-type plants. Furthermore, overexpression of MdFT1 conferred precocious flowering in apple, with altered expression of other endogenous genes, such as MdMADS12. These results suggest that MdFT1 could function to promote flowering by altering the expression of those genes and that, at least, other genes may play an important role as well in the regulation of flowering in apple. The long juvenile period of fruit trees prevents early cropping and efficient breeding. Our findings will be useful information to unveil the molecular mechanism of flowering and to develop methods to shorten the juvenile period in various fruit trees, including apple.

Four TFL1/CEN-Like Genes on Distinct Linkage Groups Show Different Expression Patterns to Regulate Vegetative and Reproductive Development in Apple (Malus×domestica Borkh.)

Recent molecular analyses in several plant species revealed that TERMINAL FLOWER1 (TFL1) and CENTRORADIALIS (CEN) homologs are involved in regulating the flowering time and/or maintaining the inflorescence meristem. In apple (Malusxdomestica Borkh.), four TFL1/CEN-like genes, MdTFL1, MdTFL1a, MdCENa and MdCENb, were found and mapped by a similar position on putatively homoeologous linkage groups. Apple TFL1/CEN-like genes functioned equivalently to TFL1 when expressed constitutively in transgenic Arabidopsis plants, suggesting that they have a potential to complement the TFL1 function. Because MdTFL1 and MdTFL1a were expressed in the vegetative tissues in both the adult and juvenile phases, they could function redundantly as a flowering repressor and a regulator of vegetative meristem identity. On the other hand, MdCENa was mainly expressed in fruit receptacles, cultured tissues and roots, suggesting that it is involved in the development of proliferating tissues but not in the control of the transition from the juvenile to the adult phase. In contrast, MdCENb was silenced in most organs probably due to gene duplication by the polyploid origin of apple. The expression patterns of MdTFL1 and MdCENa in apple were also supported by the heterologous expression of beta-glucuronidase fused with their promoter regions in transgenic Arabidopsis. Our results suggest that functional divergence of the roles in the regulation of vegetative meristem identity may have occurred among four TFL1/CEN-like genes during evolution in apple.

Apple FLOWERING LOCUS T proteins interact with transcription factors implicated in cell growth and organ development

Understanding the flowering process in apple (Malus × domestica Borkh.) is essential for developing methods to shorten the breeding period and regulate fruit yield. It is known that FLOWERING LOCUS T (FT) acts as a transmissible floral inducer in the Arabidopsis flowering network system. To clarify the molecular network of two apple FT orthologues, MdFT1 and MdFT2, we performed a yeast two-hybrid screen to identify proteins that interact with MdFT1. We identified several transcription factors, including two members of the TCP (TEOSINTE BRANCHED1, CYCLOIDEA and PROLIFERATING CELL FACTORs) family, designated MdTCP2 and MdTCP4, and an Arabidopsis thaliana VOZ1 (Vascular plant One Zinc finger protein1)-like protein, designated MdVOZ1. MdTCP2 and MdVOZ1 also interacted with MdFT2 in yeast. The expression domain of MdTCP2 and MdVOZ1 partially overlapped with that of MdFT1 and MdFT2, most strikingly in apple fruit tissue, further suggesting a potential interaction in vivo. Constitutive expression of MdTCP2, MdTCP4 and MdVOZ1 in Arabidopsis affected plant size, leaf morphology and the formation of leaf primordia on the adaxial side of cotyledons. On the other hand, chimeric MdTCP2, MdTCP4 and MdVOZ1 repressors that included the ethylene-responsive transcription factors (ERF)-associated amphiphilic repression (EAR) domain motif influenced reproduction and inflorescence architecture in transgenic Arabidopsis. These results suggest that MdFT1 and/or MdFT2 might be involved in the regulation of cellular proliferation and the formation of new tissues and that they might affect leaf and fruit development by interacting with TCP- and VOZ-family proteins. DDBJ accession nos. AB531019 (MdTCP2a mRNA), AB531020 (MdTCP2b mRNA), AB531021 (MdTCP4a mRNA), AB531022 (MdTCP4b mRNA) and AB531023 (MdVOZ1a mRNA).

Identification of QTLs for fruit quality traits in Japanese apples: QTLs for early ripening are tightly related to preharvest fruit drop

Many important apple (Malus × domestica Borkh.) fruit quality traits are regulated by multiple genes, and more information about quantitative trait loci (QTLs) for these traits is required for marker-assisted selection. In this study, we constructed genetic linkage maps of the Japanese apple cultivars ‘Orin’ and ‘Akane’ using F1 seedlings derived from a cross between these cultivars. The ‘Orin’ map consisted of 251 loci covering 17 linkage groups (LGs; total length 1095.3 cM), and the ‘Akane’ map consisted of 291 loci covering 18 LGs (total length 1098.2 cM). We performed QTL analysis for 16 important traits, and found that four QTLs related to harvest time explained about 70% of genetic variation, and these will be useful for marker-assisted selection. The QTL for early harvest time in LG15 was located very close to the QTL for preharvest fruit drop. The QTL for skin color depth was located around the position of MYB1 in LG9, which suggested that alleles harbored by ‘Akane’ are regulating red color depth with different degrees of effect. We also analyzed soluble solids and sugar component contents, and found that a QTL for soluble solids content in LG16 could be explained by the amount of sorbitol and fructose.

Genetic studies on resistance to Valsa canker in apple: genetic variance and breeding values estimated from intra- and inter-specific hybrid progeny populations

Malus sieboldii Rehd. exhibits high levels of resistance to Valsa canker caused by Valsa ceratosperma (Tode ex Fr.) Maire while cultivated apples (Malus domestica Borkh.) are susceptible to the disease. In this study, progenies from 23 full-sib families derived from both inter- and intra-specific hybridization among 16 Malus genotypes as parents were assessed for resistance to V. ceratosperma (Vc) for two seasons using an excised shoot assay to determine the pattern of inheritance of the resistance and to also estimate the variance components, narrow-sense heritability, and breeding values of parental genotypes. Generally, M. sieboldii × M. domestica and its reciprocal crosses had more resistant progenies to Vc than intra-specific crosses of M. domestica. Resistance to Vc expressed as the relative lesion length among progenies showed continuous variation irrespective of cross, suggesting the quantitative nature of the resistance to the three virulent isolates of Vc that were tested. Resistance to Vc using the progeny population was analyzed using a mixed linear model based on restricted maximum likelihood. The parental effect (general combining ability (GCA)) was significant while the interaction effect between parents (specific combining ability (SCA)) was relatively small and non-significant. The ratio of SCA/GCA variance was about 32%, suggesting that additive genetic variance had a major contribution to the total genetic variance for resistance to Vc. There was a positive correlation (r = 0.49, p < 0.01) between mid-parental GCA and SCA predictions among 23 full-sib families for the resistance. Narrow-sense heritability estimated by sib analysis was moderate

Relationships between sap flow, hydraulic conductivity, and the anatomical characteristics of stems and roots in apple rootstocks of different vigour.

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Structural and functional analyses of two new S-RNase alleles, Ssi5 and Sad5, in apple.

. The predicted breeding values (BV) of the 16 parents indicated that M. sieboldii “Sanashi 63” and “Hayanarisanashi 1” would be useful for breeding for high levels of resistance to Vc.

S-RNase genotypes of apple cultivars grown in Japan and development of a PCR-RFLP method to identify the S6- and S21-RNase alleles.

Summary We analysed the S-RNase genotypes of 23 crab apple (Malus spp.) pollinators and 102 cultivars of domestic apple (Malus pumila Mill.) by PCR amplification and digestion. Within the 23 pollinators, four pollinators, ‘Hopa’, ‘Jack’, ‘Pink Perfection’ and ‘Profusion B’, each had two unidentified S-RNase alleles. These cultivars should be useful pollinators for all domestic cultivars. Twenty-one of the domestic cultivars exhibited S-genotypes contrary to those expected from their supposed parentage, suggesting that one or both reported parents were wrong. We confirmed many of the S-RNase genotypes by pollination tests.