Abstract
We performed a comprehensive analysis of the R2R3-MYB subclass of MYB transcription factors in soybeans, focusing on GmMYB3a. Notably, we found that GmMYB3a shares high homology with MYB genes in Arabidopsis thaliana and is a MYB regulator of isoflavone biosynthesis in soybean. In our rigorous study, we have discovered that GmMYB3a exhibits nuclear localization, which concurs with its potential involvement in the biosynthesis of isoflavones. Furthermore, our analysis indicates a synergistic expression pattern between GmMYB3a and seed development, thereby strengthening the hypothesis of its critical role in the regulation of isoflavone synthesis. Transgenic experiments further demonstrated that GmMYB3a positively regulates isoflavone biosynthesis and leads to its overexpression. Moreover, GmMYB3a has been implicated in abiotic stress responses, thereby affecting soybean stress tolerance. RNA sequencing analysis revealed that GmMYB3a regulates downstream genes involved in isoflavone, flavonoid, and phenylalanine metabolism, especially the key chalcone synthase genes, CHS7 and CHS8. Moreover, GmMYB3a was shown to be tightly associated with GmCHS7 and GmCHS8 expressions, potentially regulating them directly. Yeast two-hybrid screening identified GmMYB3a interacting proteins crucial for the synthesis of physiologically active substances and abiotic stress responses. This study provided insights into the regulatory mechanisms of GmMYB3a and established a molecular network involving GmMYB3a, GmCHS7, and GmCHS8, thereby offering novel strategies for improving soybean quality and stress-tolerant breeding.