Differential proteolytic sensitivity of yeast fatty acid synthetase subunits α and β contributing to a balanced ratio of both fatty acid synthetase components

Abstract

The Saccharomyces cerevisiae genes FASI and FAS2 encoding the fl and 01 subunit of yeast fatty acid synthetase (FAS), respectively, were individually deleted by one-step gene disruption. Northern blot analysis of RNA from the resulting f a s null allele mutants indicated that deletion of FAS2 did not influence the transcription of FASI, while FASZ transcription was significantly reduced in the Afusl strain. These data suggest an activating role of subunit fl on FAS2 gene expression or, alternatively, a repression of FAS2 by an excess of its own gene product. Compared to the intact complex, the individual FAS subunits synthesized in the Afasl or Afas2 strains exhibit a considerably increased sensitivity towards the proteinases present in the yeast cell homogenate. Using yeast mutants specifically defective in the vacuolar proteinases yscA (PRAlI PEP4 gene product) and/or yscB (PRBI gene product), it was shown that in vitro, subunit LX is efficiently degraded by proteinase yscA while for degradation of subunit / 3, the combined action of proteinases yscA and yscB is necessary. In vivo, besides the vacuolar proteinases, an additional proteolytic activity specifically affecting free FAS subunit LX becomes increasingly apparent in cells entering the stationary growth phase. In contrast, under similar conditions uncomplexed FAS subunit fl is stable in strains lacking the vacuolar proteinases yscA and yscB. The reduced FAS subunit levels, at the stationary phase, were independent of the corresponding FAS transcript concentrations. Thus, differential degradation pathways are obviously removing an excess of either FAS subunit, at least under starvation conditions. A combination of both regulation of FAS gene expression and proteolysis of free FAS polypeptides may therefore explain the equimolar amounts of both FAS subunits observed in yeast wild-type cells.The biosynthesis of hetero-oligomeric proteins with defined subunit stoichiometry is presumed to be subjected to distinct control mechanisms. Other than aggregation itself, which may be achieved solely by protein self-assembly, cellular economy should require a balanced production of the individual components of an oligomer. Thereby, the accumulation of excess protein not incorporated into the aggregate would be avoided. In spite of the abundance of known protein oligomers our information on mechanisms coordinating the biosynthesis of their constituent subunits is still limited. In a few studied cases, however, such as ribosomes [I] cellular concentration of either subunit, however, does depend on the availability of the other. A large number of S. cerevisiur fk.7 mutants isolated in our laboratory simultaneously lack both subunits, although only a single F A S gene, either FASI or FAS2, is affected in each mutant [6]. The question remained open whether this effect was due to biosynthetic control or to the inherent proteolytic instability of non-aggregated FAS subunits. The recent isolation and sequence determination of both yeast FAS genes [7-101 prompted us to resume...

Microbial Type I Fatty Acid Synthases (FAS): Major Players in a Network of Cellular FAS Systems

“…Biosynthesis of the heteromultimeric fungal type I FASs requires the coordinate expression of two genes, FAS1 and FAS2, which encode subunits ␤ (FAS1) and ␣ (FAS2) of the ␣ 6 ␤ 6 FAS complex. The balanced production of the two subunits is suggested by the absence of nonaggregated FAS proteins from the yeast cell extract (28,128). In fungi other than yeast, such as Neurospora crassa and Aspergillus nidulans, the FAS1 and FAS2 genes are closely linked and arranged, in opposite orientation, around a putative common promoter.…”

A Novel Cold-Sensitive Allele of the Rate-Limiting Enzyme of Fatty Acid Synthesis, Acetyl Coenzyme A Carboxylase, Affects the Morphology of the Yeast Vacuole through Acylation of Vac8p

“…acc1 cs is defective in malonyl-CoA-dependent acyl chain elongation and the synthesis of C 26 . Supplementation with long-chain fatty acids is not sufficient to fully complement the loss of Acc1p function but is sufficient to rescue fas null alleles (13,40,42).…”

“…Acyl chains of intermediate length are elongated by a different, membrane-bound system that also uses malonylCoA for elongation (8,40,47). Null mutations in either FAS1 or FAS2 are lethal unless the cells are supplemented with exogenous fatty acids 12 to 18 carbon atoms in length (42).…”

Role of 26S proteasome and HRD genes in the degradation of 3-hydroxy-3-methylglutaryl-CoA reductase, an integral endoplasmic reticulum membrane protein.

“…Protein degradation is an essential function that has Mutant, modified, or misassembled proteins all remany roles in all eukaryotic cells (Bienkowski, 1983;quire disposal (Schuller et al, 1992;Tierney et al, 1992; Tsuji et al, 1992; Pearce and Sherman, 1995). Viral invasion strategies can include degradative tactics tCorresponding author.…”