Transcription factor Mxr1 promotes the expression of Aox1 by repressing glycerol transporter 1 in Pichia pastoris

In the methylotrophic yeast Pichia pastoris (P. pastoris), the efficient promoter of alcohol oxidase (PAox1) is induced by methanol and repressed by glycerol, but the molecular mechanism is not clear. In this study, the relationship between alcohol oxidase 1 (aox1), methanol expression regulator 1 (mxr1) and glycerol transporter 1 (gt1) was studied. By RT-PCR, it was found that the overexpression of gt1 could increase the glycerol content in cells and repress the expression of mxr1 and aox1, and the deletion of gt1 reduced the glycerol content in cells and promoted the expression of aox1. The overexpression of mxr1 could repress the expression of gt1, and the deletion of mxr1 could promote the expression of gt1 to some extent. By EMSA, Mxr1 binding sites were found in the promoter of gt1 (PGt1) (-141 to -138, CCCC), and Mxr1 could regulate the expression of gt1 by binding to PGt1. The relationships among aox1, mxr1 and gt1 revealed here provide a reference for the understanding of the mechanism of glycerol repression of PAox1.