Abstract
Related to ABSCISIC ACID INSENSITIVE3 (ABI3)/VIVIPAROUS1(VP1)(RAV) transcription factors, which encode a B3 domain and an APETALA2(AP2) domain, belong to the APETALA2/ethylene-responsive element binding factor(AP2/ERF) or B3 superfamily and play an important role in regulating plant growth and development and responding to abiotic stress. Although there have been many functional studies on RAV, the functional differences between RAVs are not clear. Therefore, in this study, the functional differences of RAVs of Medicago truncatula were analyzed. Based on sequence data from the plant transcription factor database and the M. truncatula genome database, we cloned three RAV genes from M. truncatula, named MtRAV1, MtRAV2, and MtRAV3. The cis-acting elements of these genes promoters were predicted, and the expression patterns of MtRAVs under exogenous conditions (4°C, NaCl, Polyethylene Glycol, Abscisic acid) were analyzed. MtRAVs transgenic Arabidopsis thaliana were obtained and subjected to adversity treatment. Subcellular localization results indicated that MtRAVs were located in the nucleus. A much lower expression level was observed for MtRAV3 than the levels of MtRAV1 and MtRAV2 in M. truncatula for growth in normal conditions, but under 4°C or PEG and NaCl treatment, the expression level of MtRAV3 was significantly increased. Only the MtRAV3 overexpression transgenic plants showed strong cold resistance, but the overexpressed MtRAV1 and MtRAV2 transgenic plants showed no difference from wild type plants. MtRAV transgenic plants exhibited similar response to exogenous mannitol, NaCl, and ABA, and the expression of some adverse-related marker genes were up-regulated, such as COLD REGULATED 414 THYLAKOID MEMBRANE 1 (COR414-TM1), Arabidopsis thaliana drought-induced 21 (AtDI21), and Arabidopsis thaliana phosphatidylinositol-specific phospholipase C (ATPLC). MtRAVs transgenic Arabidopsis thaliana exhibited increasing of branch number. These results indicated that there was some function redundancy during MtRAVs proteins of M. truncatula, and MtRAV3 has increased function compared to the other two genes. The results of this study should provide the foundation for future application of MtRAVs in legumes.
Highlights
RAV (Related to ABI3/VP1) transcription factors belong to the AP2 or B3 superfamilies
The survival rates of MtRAV3 transgenic plants were significantly higher than that of WT (Figure 5B). These results clearly showed that overexpression of MtRAV3 enhanced the cold tolerance of transgenic Arabidopsis thaliana and the effects on the expression of stress-responsive genes RD29B, COR78, and COR414-TM1 implied the importance of RAV protein in cold acclimation
The results indicate that the overexpression of MtRAV genes induced the expression of AtPLC1, AtPLC3, AtPLC4, AtPLC5, and ATDI21 in Arabidopsis thaliana, with the largest effect observed for the MtRAV3 transgenic plants
Summary
RAV (Related to ABI3/VP1) transcription factors belong to the AP2 or B3 superfamilies. AP2 and B3 proteins were shown to be involved in the regulation of biotic and abiotic stress responses, plant growth and development (Park et al, 2001; Zhang et al, 2005; Swaminathan et al, 2008) Transcription factors, such as AP2, DREB (CBF), ERF and LAV (ABI) families in plants, regulate the expression of defense genes in responses to biotic and abiotic stresses, seed maturation and seedling growth (Swaminathan et al, 2008; Dietz et al, 2010; Mizoi et al, 2012)
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