Celic Ivana, Verreault Alain, Boeke Jef D
High Throughput Biology Center, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Genetics. 2008 Aug;179(4):1769-84. doi: 10.1534/genetics.108.088914. Epub 2008 Jun 24.
Deacetylation of histone H3 K56, regulated by the sirtuins Hst3p and Hst4p, is critical for maintenance of genomic stability. However, the physiological consequences of a lack of H3 K56 deacetylation are poorly understood. Here we show that cells lacking Hst3p and Hst4p, in which H3 K56 is constitutively hyperacetylated, exhibit hallmarks of spontaneous DNA damage, such as activation of the checkpoint kinase Rad53p and upregulation of DNA-damage inducible genes. Consistently, hst3 hst4 cells display synthetic lethality interactions with mutations that cripple genes involved in DNA replication and DNA double-strand break (DSB) repair. In most cases, synthetic lethality depends upon hyperacetylation of H3 K56 because it can be suppressed by mutation of K56 to arginine, which mimics the nonacetylated state. We also show that hst3 hst4 phenotypes can be suppressed by overexpression of the PCNA clamp loader large subunit, Rfc1p, and by inactivation of the alternative clamp loaders CTF18, RAD24, and ELG1. Loss of CTF4, encoding a replisome component involved in sister chromatid cohesion, also suppresses hst3 hst4 phenotypes. Genetic analysis suggests that CTF4 is a part of the K56 acetylation pathway that converges on and modulates replisome function. This pathway represents an important mechanism for maintenance of genomic stability and depends upon proper regulation of H3 K56 acetylation by Hst3p and Hst4p. Our data also suggest the existence of a precarious balance between Rfc1p and the other RFC complexes and that the nonreplicative forms of RFC are strongly deleterious to cells that have genomewide and constitutive H3 K56 hyperacetylation.
由沉默调节蛋白Hst3p和Hst4p调控的组蛋白H3 K56去乙酰化作用，对于维持基因组稳定性至关重要。然而，H3 K56去乙酰化作用缺失的生理后果却鲜为人知。在此我们表明，缺乏Hst3p和Hst4p的细胞中，H3 K56持续处于高乙酰化状态，呈现出自发性DNA损伤的特征，比如检查点激酶Rad53p的激活以及DNA损伤诱导基因的上调。一致地，hst3 hst4细胞与破坏参与DNA复制和DNA双链断裂（DSB）修复基因的突变显示出合成致死相互作用。在大多数情况下，合成致死取决于H3 K56的高乙酰化状态，因为将K56突变为精氨酸可抑制这种作用，精氨酸模拟了非乙酰化状态。我们还表明，PCNA钳装载蛋白大亚基Rfc1p的过表达以及替代钳装载蛋白CTF18、RAD24和ELG1的失活，均可抑制hst3 hst4细胞的表型。编码参与姐妹染色单体黏连的复制体组分的CTF4缺失，也可抑制hst3 hst4细胞的表型。遗传分析表明，CTF4是K56乙酰化途径的一部分，该途径汇聚并调节复制体功能。此途径代表了维持基因组稳定性的重要机制，且依赖于Hst3p和Hst4p对H3 K56乙酰化的适当调控。我们的数据还表明，Rfc1p与其他RFC复合物之间存在不稳定的平衡，并且RFC的非复制形式对具有全基因组和组成性H3 K56高乙酰化的细胞具有强烈的有害作用。
