Shi X, Finkelstein A, Wolf A J, Wade P A, Burton Z F, Jaehning J A
Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver 80262, USA.
Mol Cell Biol. 1996 Feb;16(2):669-76. doi: 10.1128/MCB.16.2.669.
Regulated transcription initiation requires, in addition to RNA polymerase II and the general transcription factors, accessory factors termed mediators or adapters. We have used affinity chromatography to identify a collection of factors that associate with Saccharomyces cerevisiae RNA polymerase II (P. A. Wade, W. Werel, R. C. Fentzke, N. E. Thompson, J. F. Leykam, R. R. Burgess, J. A. Jaehning, and Z. F. Burton, submitted for publication). Here we report identification and characterization of a gene encoding one of these factors, PAF1 (for RNA polymerase-associated factor 1). PAF1 encodes a novel, highly charged protein of 445 amino acids. Disruption of PAF1 in S. cerevisiae leads to pleiotropic phenotypic traits, including slow growth, temperature sensitivity, and abnormal cell morphology. Consistent with a possible role in transcription, Paf1p is localized to the nucleus. By comparing the abundances of many yeast transcripts in isogenic wild-type and paf1 mutant strains, we have identified genes whose expression is affected by PAF1. In particular, disruption of PAF1 decreases the induction of the galactose-regulated genes three- to fivefold. In contrast, the transcript level of MAK16, an essential gene involved in cell cycle regulation, is greatly increased in the paf1 mutant strain. Paf1p may therefore be required for both positive and negative regulation of subsets of yeast genes. Like Paf1p, the GAL11 gene product is found associated with RNA polymerase II and is required for regulated expression of many yeast genes including those controlled by galactose. We have found that a gal11 paf1 double mutant has a much more severe growth defect than either of the single mutants, indicating that these two proteins may function in parallel pathways to communicate signals from regulatory factors to RNA polymerase II.
除了RNA聚合酶II和通用转录因子外，受调控的转录起始还需要被称为中介体或衔接子的辅助因子。我们利用亲和层析法鉴定了一系列与酿酒酵母RNA聚合酶II相关的因子（P.A.韦德、W.韦雷尔、R.C.芬茨克、N.E.汤普森、J.F.莱卡姆、R.R.伯吉斯、J.A.杰宁和Z.F.伯顿，已提交发表）。在此我们报告了编码其中一个因子PAF1（RNA聚合酶相关因子1）的基因的鉴定和特征。PAF1编码一种由445个氨基酸组成的新型、高度带电的蛋白质。酿酒酵母中PAF1的缺失导致多效性表型特征，包括生长缓慢、温度敏感性和异常细胞形态。与转录中的可能作用一致，Paf1p定位于细胞核。通过比较同基因野生型和paf1突变株中许多酵母转录本的丰度，我们鉴定了其表达受PAF1影响的基因。特别是，PAF1的缺失使半乳糖调控基因的诱导降低了三到五倍。相比之下，参与细胞周期调控的必需基因MAK16的转录水平在paf1突变株中大幅增加。因此，Paf1p可能是酵母基因子集正负调控所必需的。与Paf1p一样，GAL11基因产物也与RNA聚合酶II相关，并且是许多酵母基因（包括受半乳糖控制的基因）的调控表达所必需的。我们发现gal11 paf1双突变体具有比任何一个单突变体都更严重的生长缺陷，这表明这两种蛋白质可能在平行途径中发挥作用，将信号从调控因子传递给RNA聚合酶II。
