Soybean is a short-day plant, which means short day length promotes flowering. So far nine major loci, E1 to E8 and J, affecting the timing of flowering and maturity have been genetically identified in soybean. To understand the roles of soybean flowering genes in photoperiod-dependent flowering time control in soybean, we analyzed not only expression patterns of E1, E2, E3 and E4 genes as well as soybean FT homologs, including GmFT2a, GmFT5a and GmFT4, but also structural variation of E1, E2, E3, and E4 genes in various soybean accessions exhibiting a broad range of flowering time. The mRNA level of GmFT2a and GmFT5a was low in late flowering accessions, but high in late flowering accessions. In contrast, GmFT4 exhibited opposite expression pattern to those of GmFT2a and GmFT5a. Structural variation of E1, E2, E3 and E4 gene in these accessions revealed that early and moderate flowering accessions contained non-functional alleles of E1, E2, E3 and E4 genes in their genome. These results suggested that expression patterns of GmFT2a GmFT5a and GmFT4 would be important factor determining flowering time in soybean and allelic variation and genetic combination of upstream E1, E2, E3, and E4 genes would be more important in soybean flowering time control than their gene expression patterns.

Major loci controlling flowering time and maturity of short-day plant soybean, E1, E2, E3, E4, E5, E6, E7 and E8, have been identified in soybean. The gene corresponding to E2 locus is a homolog of Arabidopsis GIGANTEA (AtGI). We identified three GI homologs in soybean and are verifying their roles in day-length dependent flowering. Expression anlysis indicated that GmGIs are ubiquitously expressed at all developmental stages of soybean plants. Diurnal expression of GmGIs fluctuates within light/dark cycles of long-day (LD) and short-day (SD). GmGI2 and GmGI3 have identical expression patterns under both day length conditions with the highest peak at zeitgeber time 8 h (ZT8) under LD and at ZT4 under SD. GmGI1 shows the peak at ZT12 under LD and at ZT8 under SD. All of GmGIs exhibit the earlier peak and the shorter phase under SD than LD. The results indicated that day length affects expressions of GmGIs. Subcellular localization analysis showed that GmGIs are mainly targeted to nucleus, similar to the localization of AtGI. Overexpression of GmGIs in Arabidopsis transgenic plants showed no significant effect on flowering time nor rescue of gi-2 mutant phenotype. The results suggested that GmGIs have different molecular functions in flowering time regulation of short-day plant soybean compared to long-day plant Arabidopsis. To investigate the molecular mechanisms of GmGIs’ functions in soybean flowering time control, we intend to identify target gene of GmGIs and interacting proteins by using yeast two-hybrid assay.

FT is one of the major floral activator in photoperiod-dependent flowering pathway. To understand the role of FT homologs in flowering time control of short-day plant soybean, we identified ten soybean FT genes and named GmFTs. Phylogenetic analysis revealed that ten GmFT genes were further categorized into three subclades. Gene expression analysis showed that the most GmFT genes are mainly expressed in leaves. The expression of GmFT2a, GmFT2b, GmFT5a, and GmFT6 was strongly induced under the floral inductive short-day condition, but GmFT4 exhibited opposite expression pattern compared to those of GmFT2a, GmFT2b, GmFT5a, and GmFT6. To understand roles of GmFT genes in flowering, we generated Arabidopsis transgenic plant overexpressing GmFT genes. Both 35S:GmFT2a and 35S:GmFT5a transgenic plants showed extremely early flowering. In contrast, overexpression of GmFT4 delayed flowering of transgenic plants compared to wild type Arabidopsis. The results indicated that GmFT2a and GmFT5a might function as floral activators, while GmFT4 has an opposite function in soybean flowering. Moreover, domain swapping approaches between GmFT2a and GmFT4 revealed that the substitution of the segment B region alone, which is located in 4th exon, was sufficient to change the function of GmFT2a to floral repressor and GmFT4 to floral activator. The results suggested that soybean FT homologs have been functionally diversified during evolution and might play different roles in photoperiod-dependent flowering of soybean.

To understand molecular mechanisms underlying adaptation of plant cells to saline stress and stress memory, we developed Arabidopsis callus suspension-cultured cells adapted to high salt. Adapted cells to high salt exhibited enhanced tolerance compared to control cells. Moreover, the salt tolerance of adapted cells was stably maintained even after the stress is relieved, indicating that the salt tolerance of adapted cells was memorized. Salt-adapted and stress memorized cells were densely aggregated and formed multi-layered cell lump. Cell morphology analysis using transmission electron microscopy indicated that cell wall thickness of salt-adapted cells was significantly induced compared to control cells. In order to characterize metabolic responses of plant cells during adaptation to high salt stress as well as stress memory, we compared metabolic profiles of salt-adapted and stress-memorized cells with control cells by using NMR spectroscopy. A principle component analysis showed clear metabolic discrimination among control, salt-adapted and stress-memorized cells. Compared with control cells, metabolites related to shikimate metabolism such as tyrosine, and flavonol glycosides, which are related to protective mechanism of plant against stresses were largely up-regulated in adapted cell lines. Moreover, coniferin, a precursor of lignin, was more abundant in salt-adapted cells than control cells. The results provide new insight into metabolic level mechanisms of plant adaptation to saline stress as well as stress memory.

In order to adapt to various environmental stresses, plants have employed diverse regulatory mechanisms of gene expression. Epigenetic changes, such as DNA methylation and histone modifications play an important role in gene expression regulation under stress condition. It has been known that some of epigenetic modifications are stably inherited after mitotic and meiotic cell divisions, which is known as stress memory. To understand molecular mechanisms underlying stress memory mediated by epigenetic modifications, we developed Arabidopsis suspension-cultured cell lines adapted to high salt by stepwise increases in the NaCl concentration up to 120 mM. Adapted cell line to 120 mM NaCl, named A120, exhibited enhanced salt tolerance compared to unadapted control cells (A0). Moreover, the salt tolerance of A120 cell line was stably maintained even in the absence of added NaCl, indicating that the salt tolerance of A120 cell line was memorized even after the stress is relieved. By using salt adapted and stress memorized cell lines, we intend to analyze the changes of DNA methylation, histone modification, transcriptome, and proteome to understand molecular mechanisms underlying stress adaptation as well as stress memory in plants.

FLOWERING LOCUS T (FT) is major determinant of the length of the vegetative phase in plants. To understand the role of FT homologs in flowering time control of soybean, we identified ten soybean FT genes and named GmFTs. Expression analysis of GmFT homologs showed that the transcripts of most FT clade genes are mainly expressed in leaves. The expression of GmFT2a, GmFT2b, GmFT5a, and GmFT6 strongly induced in response to floral inductive short-day condition, but GmFT4 and GmFT6 exhibited opposite expression pattern. To understand the biological function of each GmFT/TFL1 genes in flowering time control, we ectopically expressed GmFT cDNAs in Arabidopsis under the control of CaMV 35S promoter. Interestingly, while 35S:GmFT2a and 35S:GmFT5a transgenic plant showed extremely early flowering phenotype, overexpression of GmFT4 delayed flowering. Furthermore we analyzed expression patterns GmFT genes in the leaves of Korean soybean landraces showing various flowering time. The results showed that the transcript level of two FT homologs, GmFT2a and GmFT5a, was high in early flowering landraces, but low in late flowering landraces. In contrast, GmFT4 exhibited opposite expression pattern to those of GmFT2a and GmFT5a, suggesting that GmFT4 may function antagonistically to GmFT2a and GmFT5a in flowering time control of soybean. These results demonstrated that soybean FT homologs have both unique and conserved functions in the photoperiodic control of flowering compared with those in Arabidopsis.

In order to adapt to various environmental stresses, plants have employed diverse regulatory mechanisms of gene expression. Epigenetic changes, such as DNA methylation and histone modifications play an important role in gene expression regulation under stress condition. It has been known that some of epigenetic modifications are stably inherited after mitotic and meiotic cell divisions, which is known as stress memory. To understand molecular mechanisms underlying stress memory mediated by epigenetic modifications, we developed Arabidopsis suspension-cultured cell lines adapted to high salt by stepwise increases in the NaCl concentration up to 120 mM. Adapted cell line to 120 mM NaCl, named A120, exhibited enhanced salt tolerance compared to unadapted control cells (A0). Moreover, the salt tolerance of A120 cell line was stably maintained even in the absence of added NaCl, indicating that the salt tolerance of A120 cell line was memorized even after the stress is relieved. By using salt adapted and stress memorized cell lines, we intend to analyze the changes of DNA methylation, histone modification, transcriptome, and proteome to understand molecular mechanisms underlying stress adaptation as well as stress memory in plants.