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A new regulator of proanthocyanidin (PA) biosynthesis in grapes was found by screening genes coordinately expressed with PA accumulation under different light conditions using a substantially improved method of serial analysis of gene expression (SuperSAGE). This R2R3-MYB transcription factor, VvMYBPAR, shows high protein sequence similarity with PA biosynthesis-regulating plant MYBs, such as VvMYBPA2 and TRANSPARENT TESTA2. Its transcript levels were relatively high in the skins of young berries, whereas the levels were higher in the seeds and at a maximum around veraison. In addition to its response to modified light conditions, the gene responded to abscisic acid application in the skins of cultured berries. Among the PA-specific branch genes, this transcript profile was not correlated with that of VvANR and VvLAR1 but was closely related to that of VvLAR2, suggesting different regulation of PA-specific branch genes from that of a known PA regulator, VvMYBPA2. The PA-specific regulation of VvMYBPAR was confirmed by VvMYBPAR constitutive expression in Arabidopsis in which the transgene specifically induced PA biosynthetic genes and resulted in PA accumulation in plants grown on sucrose-supplemented media to induce anthocyanin synthesis. A transient reporter assay using grapevine cells showed that VvMYBPAR activated the promoters on PA-specific branch genes and candidate genes associated with modification and transport of monomeric PA precursors, as well as the promoters of VvCHS3 and VvF3'5'Hd in the common flavonoid pathway, but not that of VvUFGT on the anthocyanin-specific branch. This new factor suggests the polygenic regulation of PA biosynthesis in grapes by closely related MYB transcription factors.

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We discovered that the origin of cT-DNA in the genome of wild-type Nicotiana glauca is the T-DNA of the mikimopine-type Ri plasmid (pRi) harbored in Agrobacterium rhizogenes. The cT-DNA was inserted into the genomic DNA of N. glauca from the position corresponding to the right border of mikimopine-type pRi. The cT-DNA contained two mikimopine synthase gene (mis) homologs, NgmisL and NgmisR, both of which were transcribed at low level in all N. glauca organs. NgMisR protein expressed in Escherichia coli has preserved Mis activity, which converts l-histidine and alpha-ketoglutaric acid to mikimopine. The mis homolog was also found in the genome of three other Nicotiana species: N. tomentosa, N. tomentosiformis, and N. tabacum; however, the site of insertion differed from that in N. glauca, suggesting that A. rhizogenes harboring mikimopine-type pRi independently infected the ancestors of some Nicotiana plants. This is the first clear evidence of a host-parasite relationship during the early evolution of Nicotiana plants. We propose that a new phylogenetic approach using opine type cT-DNA is applicable for presuming divergence in the genus Nicotiana.

SummaryThe rooting-locus gene B (rolB) on the T-DNA of the root-inducing (Ri) plasmid in Agrobacterium rhizogenes is responsible for the induction of transformed adventitious roots, although the root induction mechanism is unknown. We report here that the RolB protein of pRi1724 (1724RolB) is associated with Nicotiana tabacum 14-3-3-like protein vII (Nt14-3-3 vII) in tobacco bright yellow (BY)-2 cells. Nt14-3-3 vII directly interacts with 1724RolB protein. Green¯uorescent protein (GFP)-fused 1724RolB is localized to the nucleus. GFP-fused mutant 1724RolB proteins having a deletion or amino acid substitution are unable to interact with Nt14-3-3 vII and also show impaired nuclear localization. Moreover, these 1724RolB mutants show decreased capacity for adventitious root induction. These results suggest that adventitious root induction by 1724RolB protein correlates with its interaction with Nt14-3-3 vII and the nuclear localization of 1724RolB protein.