<i>Wrinkled</i>1 (WRI1) Homologs, AP2-Type Transcription Factors Involving Master Regulation of Seed Storage Oil Synthesis in Castor Bean (<i>Ricinus communis</i>L.)

Among APETALA2 (AP2)-type plant specific transcription factor family, WRINKLED1 (WRI1), has appeared to be a master gene transcriptionally regulating a set of carbon metabolism- and fatty acid synthesis (FAS)-related genes responsible for seed specific triacylglycerols (TAGs) storage in oil plants. B3 type transcription factors, such as ABI3 and FUS3, are known to be involved in seed development, such as seed storage protein synthesis and maturation. Based on the recent whole genome sequence data of castor bean (Ricinus communis L.), putative WRI1 homologs (RcWRI1, RcWRI2) specifically expressed in castor bean seed have been identified by comparing organ specific expression profiles among seed development-related transcription factors, seed storage specific genes (Ricin, RcOleosin) and a set of FAS genes including genes for sucrose synthase (RcSUS2), biotin carboxyl carrier protein (a subunit of acetyl-CoA carboxylase, RcBCCP2) and ketoacyl-acyl carrier protein synthase (RcKAS1). Immunoreactive signals with WRI1, FUS3 and ABI5-related polypeptides were also detected in seed specifically, consistent with the expression profiles of seed development-related genes. The WRI1 binding consensus sites, [CnTnG](n)(7)[CG], designated as the AW-box, were found at the promoter region of RcBCCP2 and RcKAS1. Thus, RcWRI1 possibly play a pivotal role in seed specific TAGs storage during seed development by directly activating FAS -related genes.

A Flower Specific Calcineurin B-Like Molecule (CBL)-Interacting Protein Kinase (CIPK) Homolog in Tomato Cultivar Micro-Tom (<i>Solanum lycopersicum</i>L.)

Floral and reproductive organs of higher plants are relatively sensitive to biotic and abiotic stresses compared with the vegetative organs. Calcineurin B-like molecule (CBL) interacting protein kinase (CIPK) has appeared to be involved in acquired tolerance and acclimation under environmental stresses such as salinity, drought and chilling. Semi-quantitative RT-PCR using the vegetative and reproductive organs of tomato Micro-Tom (Solanum lycopersicum L.) at the various developmental stages indicated that SlCIPK2 was expressed specifically in the floral organ. An anti-CIPK specific antibody recognized the recombinant SlCIPK2 specifically and cross-reacted to a CIPK-related polypeptide at a significant level in flower, particularly in stamen. The flower specific CIPK was tightly associated with the microsomes. In vitro pull-down assay of the recombinant SlCIPK2 showed that SlCIPK2 interacts with SlCBLs and stress-responsive transcription factors, SlERF7, SlCBF1 and SlAREB1. The present data suggested that the flower-specific CIPK, SlCIPK2, was involved in calcium signaling in tomato via CBLs and stress tolerance possibly mediated by the stress-responsive transcription factors in stamen.

Hsp90 and reactive oxygen species regulate thermotolerance of rice seedlings via induction of heat shock factor A2 (OsHSFA2) and galactinol synthase 1 (OsGolS1)

Heat stress induces expression of a set of thermotolerance-related genes in plants. We focused on rice (Oryza sativa L.) homologs of the gene family that encodes galactinol synthase (OsGolS), which is closely related to the Arabidopsis thaliana galactinol synthase (AtGolS) family whose expression is induced under various stresses. OsGolS1 was highly up-regulated compare to the level of OsGolS2 in re- sponse to heat stress. Interestingly, OsGolS1 was also up-regulated by treatment with the Hsp90 inhibitor, geldanamycin (GDA). Expression profiles of OsGolS1 were correlated to those of OsHsfA2 under the GDA treatments. Treatment with GDA increased expression of OsHsfA2, but marginally increased or did not change OsHsfA1 expression. Notably, gel shift assay indicated that OsHsfA2 binds directly to OsGolS1 promoter region and that OsHsfA1 also binds to the promoter regions of OsHsfA2. Both OsHsfA2 and OsGolS1 were dramatically induced in response to heat stress. Accordingly, galactinol and raffinose contents in rice seedlings increased significantly following the induction of OsGolS1. Pre-treatment of rice seedlings with raffinose or GDA improved their thermotolerance. These results suggest that OsGolS1 plays an important role in response to heat stress, possibly via the transcription cascade of OsHsfA1-OsHsfA2 that leads to galactinol and raffinose accumulation, and that the increased content of these carbohydrates is a key response factor for rice seedlings to enhance thermotolerance.

The Sucrose Starvation Signal Mediates Induction of Autophagy- and Amino Acid Catabolism-Related Genes in Cowpea Seedling

In higher plants, autophagy is bulk degradation process in vacuole necessary for survival under nutrient-limited conditions and plays important roles in senescence, development and pathogenic response, etc. Cowpea is one of the most important legume crops in semi-aride region, which is highly tolerant to drought stress. Changes of photoassimilate status by drought stress and/or sink-source balance appeared to affect autophagy and senescence of leaf in cowpea. Accordingly, we focused on roles of sucrose signal in autophagy and amino acid recycling in cowpea. Effects of starvation stress on the expression of autophagy-related genes (ATGs) and amino acid catabolism-related genes in cowpea [Vigna unguiculata (L.) Walp] were examined by Reverse transcription-polymerase chain reaction (RT-PCR) and anti-ATG8i specific antibody. Sucrose starvation stress enhanced the expression levels of VuATG8i, VuATG8c and VuATG4 incowpea seedlings. The expressions of amino acid catabolism related genes, such as asparagine synthase (VuASN1), proline dehydrogenase1 (VuProDH) and branched chain amino acid transaminase (VuBCAT2), are also up-regulated under the sucrose starvation. In contrast, high sucrose condition suppressed autophagy and the expressions of ATGs. These results indicate that sucrose starvation stress stimulates both autophagy and amino acid catabolism by regulation of ATGs and VuBCAT2. It is conceivable that sucrose starvation stress enhances autophagy in cowpea, possibly via branched chain amino acid level regulated by the starvation-induced BCAT.