Abstract
The methylotrophic yeast, Ogataea thermomethanolica TBRC656, is an attractive host organism for heterologous protein production owing to the availability of protein expression vectors and a genome-editing tool. In this study, we focused on mating-type switching and gene expression in order to elucidate its sexual life cycle and establish genetic approaches applicable for the strain. A putative mating-type gene cluster was identified in TBRC656 that is syntenic to the cluster in Ogataea parapolymorpha DL-1 (previously named Hansenula polymorpha). Like DL-1, TBRC656 possesses two mating loci, namely MATa and MAT alpha, and also shows flip-flop mating-type switching. Interestingly, unlike any other methylotrophic yeast, TBRC656 robustly switched mating type during late growth in rich medium (YPD). Under nutrient depletion, mating-type switching was observed within one hour. Transcription from both MATa and MAT alpha mating loci was detected during growth in YPD, and possibly induced upon nitrogen depletion. Gene expression from MAT alpha was detected as a single co-transcript from a three-gene array (alpha 2-alpha 1-alpha 1S). Deletion of a putative a1S ORF at the MAT alpha locus had no observed effect on mating-type switching but demonstrated significant effect on mating-type gene expression at both MATa and MAT alpha loci.
