Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia 22908.
Department of Laboratory Medicine, Jilin Medical University, 132013, China.
Genetics. 2019 May;212(1):75-91. doi: 10.1534/genetics.119.302047. Epub 2019 Mar 6.
Sir2 is a highly conserved NAD-dependent histone deacetylase that functions in heterochromatin formation and promotes replicative life span (RLS) in the budding yeast, Within the yeast rDNA locus, Sir2 is required for efficient cohesin recruitment and maintaining the stability of the tandem array. In addition to the previously reported depletion of Sir2 in replicatively aged cells, we discovered that subunits of the Sir2-containing complexes silent information regulator (SIR) and regulator of nucleolar silencing and telophase (RENT) were depleted. Several other rDNA structural protein complexes also exhibited age-related depletion, most notably the cohesin complex. We hypothesized that mitotic chromosome instability (CIN) due to cohesin depletion could be a driver of replicative aging. Chromatin immunoprecipitation assays of the residual cohesin (Mcd1-Myc) in moderately aged cells showed strong depletion from the rDNA and initial redistribution to the point centromeres, which was then lost in older cells. Despite the shift in cohesin distribution, sister chromatid cohesion was partially attenuated in aged cells and the frequency of chromosome loss was increased. This age-induced CIN was exacerbated in strains lacking Sir2 and its paralog, Hst1, but suppressed in strains that stabilize the rDNA array due to deletion of or through caloric restriction. Furthermore, ectopic expression of from a doxycycline-inducible promoter was sufficient to suppress rDNA instability in aged cells and to extend RLS. Taken together, we conclude that age-induced depletion of cohesin and multiple other nucleolar chromatin factors destabilize the rDNA locus, which then results in general CIN and aneuploidy that shortens RLS.
Sir2 是一种高度保守的 NAD 依赖性组蛋白去乙酰化酶，它在异染色质形成和促进芽殖酵母的复制寿命 (RLS) 中发挥作用。在酵母 rDNA 基因座内，Sir2 是有效募集黏连蛋白和维持串联阵列稳定性所必需的。除了先前报道的复制老化细胞中 Sir2 的耗竭之外，我们还发现 Sir2 包含的沉默信息调节因子 (SIR) 和核仁沉默和末期调节因子 (RENT) 复合物的亚基也被耗竭。其他几个 rDNA 结构蛋白复合物也表现出与年龄相关的耗竭，尤其是黏连蛋白复合物。我们假设由于黏连蛋白耗竭导致的有丝分裂染色体不稳定性 (CIN) 可能是复制衰老的驱动因素。中度老化细胞中剩余黏连蛋白 (Mcd1-Myc) 的染色质免疫沉淀分析显示，rDNA 明显耗竭，并最初重新分配到着丝粒，而在较老的细胞中则丢失。尽管黏连蛋白分布发生了转移，但姐妹染色单体的黏合在老化细胞中部分减弱，染色体丢失的频率增加。这种年龄诱导的 CIN 在缺乏 Sir2 和其同源物 Hst1 的菌株中加剧，但在由于缺失或通过热量限制稳定 rDNA 阵列的菌株中受到抑制。此外，通过四环素诱导型启动子异位表达 足以抑制老化细胞中 rDNA 的不稳定性并延长 RLS。总之，我们得出结论，年龄诱导的黏连蛋白和其他多个核仁染色质因子的耗竭使 rDNA 基因座不稳定，从而导致一般的 CIN 和非整倍体，从而缩短 RLS。
