Soybean Calmodulin-Binding Transcription Activators, GmCAMTA2 and GmCAMTA8, Coordinate the Circadian Regulation of Developmental Processes and Drought Stress Responses.

The calmodulin-binding transcription activators (CAMTAs) mediate transcriptional regulation of development, growth, and responses to various environmental stresses in plants. To understand the biological roles of soybean CAMTA (GmCAMTA) family members in response to abiotic stresses, we characterized expression patterns of 15 GmCAMTA genes in response to various abiotic stresses. The GmCAMTA genes exhibited distinct circadian regulation expression patterns and were differently expressed in response to salt, drought, and cold stresses. Interestingly, the expression levels of GmCAMTA2, GmCAMTA8, and GmCAMTA12 were higher in stem tissue than in other soybean tissues. To determine the roles of GmCAMTAs in the regulation of developmental processes and stress responses, we isolated GmCAMTA2 and GmCAMTA8 cDNAs from soybean and generated Arabidopsis overexpressing transgenic plants. The GmCAMTA2-OX and GmCAMTA8-OX plants showed hypersensitivity to drought stress. The water in the leaves of GmCAMTA2-OX and GmCAMTA8-OX plants was lost faster than that in wild-type (WT) plants under drought-stress conditions. In addition, stress-responsive genes were down-regulated in the GmCAMTA2-OX and GmCAMTA8-OX plants under drought stress conditions compared to WT plants. Our results suggest that GmCAMTA2 and GmCAMTA8 genes are regulated by circadian rhythms and function as negative regulators in development and drought stress responses.

High-Temperature Conditions Promote Soybean Flowering through the Transcriptional Reprograming of Flowering Genes in the Photoperiod Pathway.

Global warming has an impact on crop growth and development. Flowering time is particularly sensitive to environmental factors such as day length and temperature. In this study, we investigated the effects of global warming on flowering using an open-top Climatron chamber, which has a higher temperature and CO2 concentration than in the field. Two different soybean cultivars, Williams 82 and IT153414, which exhibited different flowering times, were promoted flowering in the open-top Climatron chamber than in the field. We more specifically examined the expression patterns of soybean flowering genes on the molecular level under high-temperature conditions. The elevated temperature induced the expression of soybean floral activators, GmFT2a and GmFT5a as well as a set of GmCOL genes. In contrast, it suppressed floral repressors, E1 and E2 homologs. Moreover, high-temperature conditions affected the expression of these flowering genes in a day length-independent manner. Taken together, our data suggest that soybean plants properly respond and adapt to changing environments by modulating the expression of a set of flowering genes in the photoperiod pathway for the successful production of seeds and offspring.

Microtubule Dynamics Plays a Vital Role in Plant Adaptation and Tolerance to Salt Stress.

Although recent studies suggest that the plant cytoskeleton is associated with plant stress responses, such as salt, cold, and drought, the molecular mechanism underlying microtubule function in plant salt stress response remains unclear. We performed a comparative proteomic analysis between control suspension-cultured cells (A0) and salt-adapted cells (A120) established from Arabidopsis root callus to investigate plant adaptation mechanisms to long-term salt stress. We identified 50 differentially expressed proteins (45 up- and 5 down-regulated proteins) in A120 cells compared with A0 cells. Gene ontology enrichment and protein network analyses indicated that differentially expressed proteins in A120 cells were strongly associated with cell structure-associated clusters, including cytoskeleton and cell wall biogenesis. Gene expression analysis revealed that expressions of cytoskeleton-related genes, such as FBA8, TUB3, TUB4, TUB7, TUB9, and ACT7, and a cell wall biogenesis-related gene, CCoAOMT1, were induced in salt-adapted A120 cells. Moreover, the loss-of-function mutant of Arabidopsis TUB9 gene, tub9, showed a hypersensitive phenotype to salt stress. Consistent overexpression of Arabidopsis TUB9 gene in rice transgenic plants enhanced tolerance to salt stress. Our results suggest that microtubules play crucial roles in plant adaptation and tolerance to salt stress. The modulation of microtubule-related gene expression can be an effective strategy for developing salt-tolerant crops.

Metabolic Adjustment of Arabidopsis Root Suspension Cells During Adaptation to Salt Stress and Mitotic Stress Memory.

Sessile plants reprogram their metabolic and developmental processes during adaptation to prolonged environmental stresses. To understand the molecular mechanisms underlying adaptation of plant cells to saline stress, we established callus suspension cell cultures from Arabidopsis roots adapted to high salt for an extended period of time. Adapted cells exhibit enhanced salt tolerance compared with control cells. Moreover, acquired salt tolerance is maintained even after the stress is relieved, indicating the existence of a memory of acquired salt tolerance during mitotic cell divisions, known as mitotic stress memory. Metabolite profiling using 1H-nuclear magnetic resonance (NMR) spectroscopy revealed metabolic discrimination between control, salt-adapted and stress-memory cells. Compared with control cells, salt-adapted cells accumulated higher levels of sugars, amino acids and intermediary metabolites in the shikimate pathway, such as coniferin. Moreover, adapted cells acquired thicker cell walls with higher lignin contents, suggesting the importance of adjustments of physical properties during adaptation to elevated saline conditions. When stress-memory cells were reverted to normal growth conditions, the levels of metabolites again readjusted. Whereas most of the metabolic changes reverted to levels intermediate between salt-adapted and control cells, the amounts of sugars, alanine, γ-aminobutyric acid and acetate further increased in stress-memory cells, supporting a view of their roles in mitotic stress memory. Our results provide insights into the metabolic adjustment of plant root cells during adaptation to saline conditions as well as pointing to the function of mitotic memory in acquired salt tolerance.

Arabidopsis CCoAOMT1 Plays a Role in Drought Stress Response via ROS- and ABA-Dependent Manners.

Plants possess adaptive reprogramed modules to prolonged environmental stresses, including adjustment of metabolism and gene expression for physiological and morphological adaptation. CCoAOMT1 encodes a caffeoyl CoA O-methyltransferase and is known to play an important role in adaptation of Arabidopsis plants to prolonged saline stress. In this study, we showed that the CCoAOMT1 gene plays a role in drought stress response. Transcript of CCoAOMT1 was induced by salt, dehydration (drought), and methyl viologen (MV), and loss of function mutants of CCoAOMT1, ccoaomt1-1, and ccoaomt1-2 exhibit hypersensitive phenotypes to drought and MV stresses. The ccoaomt1 mutants accumulated higher level of H2O2 in the leaves and expressed lower levels of drought-responsive genes including RD29B, RD20, RD29A, and ERD1, as well as ABA3 3 and NCED3 encoding ABA biosynthesis enzymes during drought stress compared to wild-type plants. A seed germination assay of ccoaomt1 mutants in the presence of ABA also revealed that CCoAOMT1 functions in ABA response. Our data suggests that CCoAOMT1 plays a positive role in response to drought stress response by regulating H2O2 accumulation and ABA signaling.

Diversification in Functions and Expressions of Soybean FLOWERING LOCUS T Genes Fine-Tunes Seasonal Flowering.

The proper timing of flowering in response to environmental changes is critical for ensuring crop yields. FLOWERING LOCUS T (FT) homologs of the phosphatidylethanolamine-binding protein family play important roles as floral integrators in many crops. In soybean, we identified 17 genes of this family, and characterized biological functions in flowering for ten FT homologs. Overexpression of GmFT homologs in Arabidopsis revealed that a set of GmFT homologs, including GmFT2a/2b, GmFT3a/3b, and GmFT5a/5b, promoted flowering similar to FT; in contrast, GmFT1a/1b, GmFT4, and GmFT6 delayed flowering. Consistently, expressions of GmFT2a, GmFT2b, and GmFT5a were induced in soybean leaves in response to floral inductive short days, whereas expressions of GmFT1a and GmFT4 were induced in response to long days. Exon swapping analysis between floral activator GmFT2a and floral repressor GmFT4 revealed that the segment B region in the fourth exon is critical for their antagonistic functions. Finally, expression analysis of GmFT2a, GmFT5a, and GmFT4 in soybean accessions exhibiting various flowering times indicated that the mRNA levels of GmFT2a and GmFT5a were higher in early flowering accessions than in late-flowering accessions, while GmFT4 showed the opposite pattern. Moreover, the relative mRNA levels between GmFT2a/GmFT5a and GmFT4 was important in determining day length-dependent flowering in soybean accessions. Taken together, our results suggest that the functions of GmFT homologs have diversified into floral activators and floral repressors during soybean evolution, and the timing of flowering in response to changing day length is determined by modulating the activities of antagonistic GmFT homologs.

Lignin biosynthesis genes play critical roles in the adaptation of Arabidopsis plants to high-salt stress.

Salinity is a major abiotic stressor that limits the growth, development, and reproduction of plants. Our previous metabolic analysis of high salt-adapted callus suspension cell cultures from Arabidopsis roots indicated that physical reinforcement of the cell wall is an important step in adaptation to saline conditions. Compared to normal cells, salt-adapted cells exhibit an increased lignin content and thickened cell wall. In this study, we investigated not only the lignin biosynthesis gene expression patterns in salt-adapted cells, but also the effects of a loss-of-function of CCoAOMT1, which plays a critical role in the lignin biosynthesis pathway, on plant responses to high-salt stress. Quantitative real-time PCR analysis revealed higher mRNA levels of genes involved in lignin biosynthesis, including CCoAOMT1, 4CL1, 4CL2, COMT, PAL1, PAL2, and AtPrx52, in salt-adapted cells relative to normal cells, which suggests activation of the lignin biosynthesis pathway in salt-adapted cells. Moreover, plants harboring the CCoAOMT1 mutants, ccoaomt1-1 and ccoaomt1-2, were phenotypically hypersensitive to salt stress. Our study has provided molecular and genetic evidence indicating the importance of enhanced lignin accumulation in the plant cell wall during the responses to salt stress.