Wright Robin, Parrish Mark L, Cadera Emily, Larson Lynnelle, Matson Clinton K, Garrett-Engele Philip, Armour Chris, Lum Pek Yee, Shoemaker Daniel D
University of Minnesota, Department of Genetics, Cell Biology and Development, 321 Church Street, 6-160 Jackson Hall, Minneapolis, MN 55455, USA.
Yeast. 2003 Jul 30;20(10):881-92. doi: 10.1002/yea.994.
Increased levels of HMG-CoA reductase induce cell type- and isozyme-specific proliferation of the endoplasmic reticulum. In yeast, the ER proliferations induced by Hmg1p consist of nuclear-associated stacks of smooth ER membranes known as karmellae. To identify genes required for karmellae assembly, we compared the composition of populations of homozygous diploid S. cerevisiae deletion mutants following 20 generations of growth with and without karmellae. Using an initial population of 1,557 deletion mutants, 120 potential mutants were identified as a result of three independent experiments. Each experiment produced a largely non-overlapping set of potential mutants, suggesting that differences in specific growth conditions could be used to maximize the comprehensiveness of similar parallel analysis screens. Only two genes, UBC7 and YAL011W, were identified in all three experiments. Subsequent analysis of individual mutant strains confirmed that each experiment was identifying valid mutations, based on the mutant's sensitivity to elevated HMG-CoA reductase and inability to assemble normal karmellae. The largest class of HMG-CoA reductase-sensitive mutations was a subset of genes that are involved in chromatin structure and transcriptional regulation, suggesting that karmellae assembly requires changes in transcription or that the presence of karmellae may interfere with normal transcriptional regulation.
HMG-CoA还原酶水平的升高会诱导内质网发生细胞类型和同工酶特异性增殖。在酵母中，由Hmg1p诱导的内质网增殖由与核相关的光滑内质网膜堆叠组成，称为卡氏小体。为了鉴定卡氏小体组装所需的基因，我们比较了纯合二倍体酿酒酵母缺失突变体群体在有和没有卡氏小体的情况下生长20代后的组成。使用1557个缺失突变体的初始群体，经过三个独立实验，鉴定出120个潜在突变体。每个实验产生的潜在突变体集合在很大程度上不重叠，这表明特定生长条件的差异可用于最大化类似平行分析筛选的全面性。在所有三个实验中仅鉴定出两个基因，即UBC7和YAL011W。随后对单个突变菌株的分析证实，基于突变体对升高的HMG-CoA还原酶的敏感性以及无法组装正常卡氏小体，每个实验都鉴定出了有效的突变。对HMG-CoA还原酶敏感的最大一类突变是参与染色质结构和转录调控的基因子集，这表明卡氏小体组装需要转录变化，或者卡氏小体的存在可能会干扰正常的转录调控。
