Schüller H J, Förtsch B, Rautenstrauss B, Wolf D H, Schweizer E
Lehrstuhl für Biochemie, Universität Erlangen-Nürnberg, Federal Republic of Germany.
Eur J Biochem. 1992 Feb 1;203(3):607-14. doi: 10.1111/j.1432-1033.1992.tb16590.x.
The Saccharomyces cerevisiae genes FAS1 and FAS2 encoding the beta and alpha subunit of yeast fatty acid synthetase (FAS), respectively, were individually deleted by one-step gene disruption. Northern blot analysis of RNA from the resulting fas null allele mutants indicated that deletion of FAS2 did not influence the transcription of FAS1, while FAS2 transcription was significantly reduced in the delta fas1 strain. These data suggest an activating role of subunit beta on FAS2 gene expression or, alternatively, a repression of FAS2 by an excess of its own gene product. Compared to the intact alpha 6 beta 6 complex, the individual FAS subunits synthesized in the delta fas1 or delta fas2 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 (PRA1/ PEP4 gene product) and/or yscB (PRB1 gene product), it was shown that in vitro, subunit alpha is efficiently degraded by proteinase yscA while for degradation of subunit beta, 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 alpha becomes increasingly apparent in cells entering the stationary growth phase. In contrast, under similar conditions uncomplexed FAS subunit beta 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.
酿酒酵母中分别编码酵母脂肪酸合成酶（FAS）β亚基和α亚基的基因FAS1和FAS2，通过一步基因破坏被单独缺失。对所得fas无效等位基因突变体的RNA进行Northern印迹分析表明，FAS2的缺失不影响FAS1的转录，而在Δfas1菌株中FAS2转录显著降低。这些数据表明β亚基对FAS2基因表达具有激活作用，或者其自身基因产物过量会对FAS2产生抑制作用。与完整的α6β6复合物相比，在Δfas1或Δfas2菌株中合成的单个FAS亚基对酵母细胞匀浆中存在的蛋白酶表现出显著增加的敏感性。使用在液泡蛋白酶yscA（PRA1/PEP4基因产物）和/或yscB（PRB1基因产物）中存在特异性缺陷的酵母突变体，结果表明在体外，α亚基被蛋白酶yscA有效降解，而β亚基的降解则需要蛋白酶yscA和yscB的共同作用。在体内，除了液泡蛋白酶外，在进入稳定生长期的细胞中，一种特异性影响游离FAS亚基α的额外蛋白水解活性变得越来越明显。相反，在类似条件下，未复合的FAS亚基β在缺乏液泡蛋白酶yscA和yscB的菌株中是稳定的。在稳定期，FAS亚基水平的降低与相应的FAS转录本浓度无关。因此，至少在饥饿条件下，不同的降解途径显然在去除过量的任何一种FAS亚基。因此，FAS基因表达的调控和游离FAS多肽的蛋白水解作用相结合，可能解释了在酵母野生型细胞中观察到的两种FAS亚基等摩尔量的现象。
