The transcription factor Snf1p is involved in a Tup1p-independent manner in the glucose regulation of the major methanol metabolism genes of Hansenula polymorpha

Abstract

Hansenula polymorpha is a methylotrophic yeast widely employed in biotechnology as a "protein factory". Most promoters used for heterologous protein expression, like MOX (methanol oxidase) and DAS (di-hydroxy acetone synthase), are involved in the peroxisomal methanol metabolism (C 1 metabolism) and are under strong glucose repression. Interestingly, the MOX promoter is subjected to glucose regulation also in Saccharomyces cerevisiae, a non-methylotrophic yeast in which this phenomenon is well studied. In this species, the transcription factor Tup1p plays an essential role in glucose repression of several genes. This effect is counteracted by the activator Snf1p when glucose is exhausted from medium. Therefore, to test whether this regulatory circuit has been conserved in H. polymorpha, HpTUP1 and HpSNF1 were partially cloned and disrupted. Deletion of HpTUP1 did not affect glucose repression of the major C 1 metabolism genes (MOX, DAS). Thus, though conserved, HpTUP1 does not seem to take part in a general glucose repression in H. polymorpha. In contrast, the deletion of HpSNF1 led to significant decreases in the activation of these genes in the absence of glucose. Therefore, the effect of HpSNF1 in transcriptional activation may be through an HpTUP1-independent circuit.

The role ofHansenula polymorpha MIG1homologues in catabolite repression and pexophagy

“…However, Δ mig1 and Δ mig1 Δ mig2 mutants could grow on methanol plates in the presence of 2‐deoxyglucose, suggesting an impairment of glucose repression. The H. polymorpha Δ tup1 mutant, which is not disturbed for glucose repression (Oliveira et al , 2003), was used as a control.…”

“…However, Tup1 is involved in pleiotropic functions through interaction with specific DNA‐binding proteins for each functionally related set of genes (reviewed in Malave & Dent, 2006), while the Mig1‐dependent regulation is thought to be more specific, with a limited set of the target genes that include those repressed by glucose (Murad et al , 2001). Therefore, our results are in line with the previous observation that HpTup1 is not required for glucose repression of peroxisomal enzymes (Oliveira et al , 2003), and suggest that, at least for alcohol oxidase, operation of the classical S. cerevisiae pathway with Mig1/2‐mediated Tup1‐Ssn6 binding to the repressible promoter is unlikely. However, the redundant function of these repressors cannot be excluded, and the effect of combining all three mutations in one strain would be interesting to examine.…”

“…The above findings suggested that glucose‐repressible genes of C‐1 metabolism in H. polymorpha may be the target of Mig1/(Tup1‐Ssn6)‐mediated transcriptional repression, analogous to the case of baker's yeast. However, Oliveira et al (2003) reported that HpTup1 is not essential for glucose repression of peroxisomal enzymes in H. polymorpha .…”

Untitled

“…In contrast to the TUP1 knockout where no apparent phenotype could be observed, transcription of the MOX and DAS genes was reduced under methanol inducing conditions in the snf1Δ strain. Interestingly, although the mRNA levels of these two genes were significantly reduced, the growth of the snf1Δ was not distinguishable from the wild type strain, indicating that reduction of the MOX and DAS transcription does not result in a slowdown or bottleneck in methanol utilisation [133]. …”

Trm2p-dependent derepression is essential for methanol-specific gene activation in the methylotrophic yeast Candida boidinii

“…Snf1p regulates the binding of Adr1p to chromatin in response to derepression. In H. polymorpha , disruption of the SNF1 homologue results in reduced transcription of MOX and DAS genes in cells grown on methanol, although the rate of growth on methanol is indistinguishable from that of the wild‐type strain (Oliveira et al , 2003). It is likely that the Snf1p homologue also regulates the binding of Trm2p to chromatin in C. boidinii .…”