Molecular characterization of Candida boidinii MIG1 and its role in the regulation of methanol‐inducible gene expression

Abstract

Methanol-inducible gene promoters in methanol-utilizing yeasts are used in high-level heterologous gene expression systems. Generally, expression of methanol-inducible genes is completely repressed by the presence of glucose. In this study we identified the MIG1 gene in Candida boidinii, which encodes a homologue of the glucose repressor Mig1p of Saccharomyces cerevisiae. Disruption of the CbMIG1 gene had no growth effect on various carbon sources. Activation of the methanol-inducible AOD1 gene, which encodes alcohol oxidase, was increased in the early stage of methanol induction when cells of the CbMIG1-disrupted strain were transferred from glucose medium to methanol medium. Furthermore, CbMig1p tagged with yellow fluorescent protein was primarily localized in the nucleus of glucose-grown cells, but was diffuse in the cytosol of methanol-grown cells. This cytosolic diffusion in methanol-grown cells occurred in a CbMsn5p-dependent manner. These results suggest that CbMig1p is involved in negative regulation of methanol-inducible gene expression in C. boidinii.

Molecular Characterization of Hap Complex Components Responsible for Methanol-Inducible Gene Expression in the Methylotrophic Yeast Candida boidinii

“…The key enzymes of methanol metabolism in methylotrophic yeasts are highly induced by methanol and are virtually absent when cells are growing on glucose. We have studied the regulation of the methanol-inducible genes necessary for methanol metabolism and characterized several transcription factors involved in their regulation in C. boidinii (8)(9)(10)(11). Based on our previous results, we have proposed three modes of regulation during methanol-inducible gene expression, that is, CbMig1p- dependent glucose repression, CbTrm2p-dependent derepression, and CbTrm1p-dependent methanol-specific induction.…”

“…In previous studies, we identified several transcription factors responsible for methanol-inducible gene expression in C. boidinii and revealed the specific function of each transcription factor, i.e., CbMig1p in glucose repression, CbTrm2p in glucose derepression, and CbTrm1p in methanol-specific induction (8)(9)(10)(11). Among these three modes of regulation, the molecular mechanism of glucose repression/derepression has been extensively studied in the budding yeast Saccharomyces cerevisiae after a shift of the carbon source from glucose to the nonfermentable carbon source ethanol or glycerol (12,13).…”

Methanol independent induction in Pichia pastoris by simple derepressed overexpression of single transcription factors

“…This fact enables the use of unmodified host strains without additional deletion of typical repressors ( MIG1 and MIG2 , S 2). Since Mig1/2p homologs in S. cerevisiae and related methylotrophic yeasts (Stasyk et al, ; Zhai, Yurimoto, & Sakai, ) release glucose repression upon glucose depletion, such knockouts appear to become obsolete under derepressed conditions in P. pastoris . Once derepressed conditions are reached, overexpression of the factors appears to mimic methanol induction.…”

“…This fact enables the use of unmodified host strains without additional deletion of typical repressors (MIG1 and MIG2, S 2). Since Mig1/2p homologs in S. cerevisiae and related methylotrophic yeasts (Stasyk et al, 2007;Zhai, Yurimoto, & Sakai, 2012)…”

Unique C-terminal region of Hap3 is required for methanol-regulated gene expression in the methylotrophic yeast Candida boidinii

“…Exhaustion of glucose releases glucose repression, resulting in activation of methanol-regulated genes by CbTrm2p, which does not require methanol for gene activation (derepression). In addition, the presence of methanol induces maximum activation of methanolregulated genes via CbTrm1p (methanol induction; methanol-specific induction) (Hartner & Glieder, 2006;Sasano et al, 2008;Yurimoto, 2009;Sasano et al, 2010;Zhai et al, 2012).…”