Research on the Mechanism of Mating-Type-Specific Gene Expression by Transcription Factor Mata2 in Miso and Soy Sauce...

The mating-type locus of the miso and soy sauce yeast Zygosaccharomyces sp. contains a gene encoding the transcription factor Mata2 (hereafter ZygoMata2), which has a DNA-binding domain containing a high mobility group (HMG)-box not found in the baker's yeast Saccharomyces cerevisiae. To investigate the role of ZygoMata2 in mating-type a expression, we constructed mutant strains of ZygoMATa2 using CRISPR-Cas9. The resulting mutants showed reduced expression of ZygoSTE6, which is specific to mating-type a, and did not mate, suggesting that ZygoMata2 regulates the expression of mating-type a in Zygosaccharomyces sp. Expression of ZygoSTE6 was observed when the HMG-box of ZygoMata2 was exchanged with that of Mat1-Mc, which regulates mating-type expression in Schizosaccharomyces pombe, indicating that ZygoMata2 recognizes its target DNA sequence via the HMG-box. Substitution of the predicted upstream activation sequence (UAS) in ZygoSTE6 with the UAS of a mating-type a-specific gene from another yeast species led to mating-type a-specific gene expression in transformed Zygosaccharomyces sp. yeast, indicating that ZygoMata2 can recognize the UAS of mating-type a-specific genes of other species. We speculated that the flexibility in the binding sequences of ZygoMata2 is due to synergistic or cooperative binding with the DNA-binding protein ZygoMcm1. This hypothesis was supported by replacement of the UAS of ZygoSTE6 with the complete palindrome sequence, P(PAL), and by the results of protein and DNA structure prediction using AlphaFold. Collectively, our study indicates that ZygoMata2 regulates mating-type a-specific gene expression via its HMG-box, which shows flexible recognition of the binding sequence of other HMG-box transcription factors.