Transcriptional regulation of sterol metabolism in Saccharomyces cerevisiae

Abstract

Sterols are important in the physiology of all eukaryotes for membrane fluidity and function. Yeast sterols regulate endogenous sterol biosynthesis by a transcriptional feedback mechanism. The ergosterol biosynthetic pathway in the budding yeast S. cerevisiae is highly conserved with the cholesterol biosynthetic pathway in mammals. Erg26 (4[alpha]-carboxysterol-C3 dehydrogenase) along with Erg25 (sterol-c-4-methyl-oxidase) and Erg27 (c-3-keto-reductase) is required for conversion of 4,4-dimethylzymosterol to zymosterol. erg26-1ts cells harbor defects in the 4[alpha]-carboxysterol-C3 dehydrogenase activity necessary for conversion of 4,4-dimethylzymosterol to zymosterol. A genetic screen aimed at cloning recessive mutations remediating the temperature-sensitive growth defect of erg26-1ts cells has resulted in the isolation of recessive alleles of ERG1 and ERG7, designated erg1-1 and erg7-1. We have used these and other erg mutants to uncover novel yeast sterol transcriptional and post translational coordinate regulatory pathways. We demonstrate that the expression of multiple ERG genes are coordinately transcriptionally regulated in response to blocks in sterol biosynthesis. Transcriptional regulation requires the transcription factors Upc2p and Ecm22p. Curiously, the ERG25 and ERG1 genes contain putative promoter SRE sites, while ERG26, ERG27, and ERG7 genes do not contain any known putative SRE sites. Next, we addressed the potential physiological role of SREs in sterol-dependent transcriptional regulation. Our deletion analysis and site-directed mutagenesis studies indicate the SREs within ERG25 and ERG3 are not required for sterol-dependent regulation, while the single SRE site in ERG1 is required for sterol-dependent regulation. Interestingly, Upc2 and Ecm22 are still required for regulation ERG25 and ERG3 gene expression in the absence of their SREs and ERG1 gene expression in the presence of its single SRE site. To define how this novel SRE-independent pathway is regulated, we have examined the transcriptional regulation of UPC2 and ECM22 in response to sterol deprivation. We find that the sterol-dependent up regulation of UPC2, not ECM22 occurs concomitant with ERG up regulation. UPC2 does not contain any putative SRE sites and we have narrowed down the promoter region within UPC2 that is sterol-responsive. Our results suggest a novel SRE-independent mechanism regulating ERG25 and ERG3, and an SRE-dependent mechanism regulating ERG1 gene expression that remains Upc2/Ecm22 dependent. We propose a novel transcriptional signaling pathway, which coordinately regulates the transcription of ERG genes at multiple points in the sterol pathway. The pathway uses SRE-dependent and SRE-independent mechanisms, and requires the Upc2 and Ecm22 transcription factors, thus tying in multiple cell events to the single process of transcriptional activation in response to sterol deprivation.