Smith J S, Brachmann C B, Pillus L, Boeke J D
Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Genetics. 1998 Jul;149(3):1205-19. doi: 10.1093/genetics/149.3.1205.
Transcriptional silencing in Saccharomyces cerevisiae occurs at the silent mating-type loci HML and HMR, at telomeres, and at the ribosomal DNA (rDNA) locus RDN1. Silencing in the rDNA occurs by a novel mechanism that depends on a single Silent Information Regulator (SIR) gene, SIR2. SIR4, essential for other silenced loci, paradoxically inhibits rDNA silencing. In this study, we elucidate a regulatory mechanism for rDNA silencing based on the finding that rDNA silencing strength directly correlates with cellular Sir2 protein levels. The endogenous level of Sir2p was shown to be limiting for rDNA silencing. Furthermore, small changes in Sir2p levels altered rDNA silencing strength. In rDNA silencing phenotypes, sir2 mutations were shown to be epistatic to sir4 mutations, indicating that SIR4 inhibition of rDNA silencing is mediated through SIR2. Furthermore, rDNA silencing is insensitive to SIR3 overexpression, but is severely reduced by overexpression of full-length Sir4p or a fragment of Sir4p that interacts with Sir2p. This negative effect of SIR4 overexpression was overridden by co-overexpression of SIR2, suggesting that SIR4 directly inhibits the rDNA silencing function of SIR2. Finally, genetic manipulations of SIR4 previously shown to promote extended life span also resulted in enhanced rDNA silencing. We propose a simple model in which telomeres act as regulators of rDNA silencing by competing for limiting amounts of Sir2 protein.
酿酒酵母中的转录沉默发生在沉默交配型基因座HML和HMR、端粒以及核糖体DNA（rDNA）基因座RDN1处。rDNA中的沉默通过一种依赖于单个沉默信息调节因子（SIR）基因SIR2的新机制发生。对其他沉默基因座至关重要的SIR4却反常地抑制rDNA沉默。在本研究中，基于rDNA沉默强度与细胞Sir2蛋白水平直接相关这一发现，我们阐明了rDNA沉默的调控机制。结果表明，Sir2p的内源性水平对rDNA沉默具有限制作用。此外，Sir2p水平的微小变化会改变rDNA沉默强度。在rDNA沉默表型中，sir2突变对sir4突变呈上位性，表明SIR4对rDNA沉默的抑制作用是通过SIR2介导的。此外，rDNA沉默对SIR3的过表达不敏感，但全长Sir4p或与Sir2p相互作用的Sir4p片段的过表达会使其严重降低。SIR2的共过表达克服了SIR4过表达的这种负面影响，表明SIR4直接抑制SIR2的rDNA沉默功能。最后，先前显示可促进延长寿命的SIR4基因操作也导致rDNA沉默增强。我们提出了一个简单的模型，其中端粒通过竞争有限量的Sir2蛋白来充当rDNA沉默的调节因子。
