Ye Yanfang, Kirkham-McCarthy Lucy, Lahue Robert S
Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland, Galway, University Road, Galway, Ireland; College of Crop Science, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland, Galway, University Road, Galway, Ireland; NCBES Galway Neuroscience Centre, School of Natural Sciences, National University of Ireland, Galway University Road, Galway, Ireland.
DNA Repair (Amst). 2016 Jul;43:1-8. doi: 10.1016/j.dnarep.2016.04.012. Epub 2016 May 2.
Trinucleotide repeats (TNRs) are tandem arrays of three nucleotides that can expand in length to cause at least 17 inherited human diseases. Somatic expansions in patients can occur in differentiated tissues where DNA replication is limited and cannot be a primary source of somatic mutation. Instead, mouse models of TNR diseases have shown that both inherited and somatic expansions can be suppressed by the loss of certain DNA repair factors. It is generally believed that these repair factors cause misprocessing of TNRs, leading to expansions. Here we extend this idea to show that the Mre11-Rad50-Xrs2 (MRX) complex of Saccharomyces cerevisiae is a causative factor in expansions of short TNRs. Mutations that eliminate MRX subunits led to significant suppression of expansions whereas mutations that inactivate Rad51 had only a minor effect. Coupled with previous evidence, this suggests that MRX drives expansions of short TNRs through a process distinct from homologous recombination. The nuclease function of Mre11 was dispensable for expansions, suggesting that expansions do not occur by Mre11-dependent nucleolytic processing of the TNR. Epistasis between MRX and post-replication repair (PRR) was tested. PRR protects against expansions, so a rad5 mutant gave a high expansion rate. In contrast, the mre11 rad5 double mutant gave a suppressed expansion rate, indistinguishable from the mre11 single mutant. This suggests that MRX creates a TNR substrate for PRR. Protein acetylation was also tested as a mechanism regulating MRX activity in expansions. Six acetylation sites were identified in Rad50. Mutation of all six lysine residues to arginine gave partial bypass of a sin3 HDAC mutant, suggesting that Rad50 acetylation is functionally important for Sin3-mediated expansions. Overall we conclude that yeast MRX helps drive expansions of short TNRs by a mechanism distinct from its role in homologous recombination and independent of the nuclease function of Mre11.
三核苷酸重复序列（TNRs）是由三个核苷酸组成的串联阵列，其长度可扩展，从而导致至少17种人类遗传性疾病。患者体内的体细胞扩增可发生在DNA复制受限的分化组织中，且不能成为体细胞突变的主要来源。相反，TNR疾病的小鼠模型表明，遗传性和体细胞扩增都可因某些DNA修复因子的缺失而受到抑制。人们普遍认为，这些修复因子会导致TNRs加工错误，从而导致扩增。在此，我们拓展了这一观点，以表明酿酒酵母的Mre11-Rad50-Xrs2（MRX）复合物是短TNRs扩增的致病因素。消除MRX亚基的突变导致扩增显著受到抑制，而使Rad51失活的突变只有轻微影响。结合先前的证据，这表明MRX通过一个不同于同源重组的过程驱动短TNRs的扩增。Mre11的核酸酶功能对于扩增是可有可无的，这表明扩增并非通过Mre11依赖的TNR核酸水解加工发生。对MRX与复制后修复（PRR）之间的上位性进行了测试。PRR可防止扩增，因此rad5突变体具有较高的扩增率。相比之下，mre11 rad5双突变体的扩增率受到抑制，与mre11单突变体无明显差异。这表明MRX为PRR创造了一个TNR底物。还测试了蛋白质乙酰化作为调节MRX在扩增中活性的一种机制。在Rad50中鉴定出六个乙酰化位点。将所有六个赖氨酸残基突变为精氨酸可部分绕过sin3 HDAC突变体，这表明Rad50乙酰化对于Sin3介导的扩增在功能上很重要。总体而言，我们得出结论，酵母MRX通过一种不同于其在同源重组中的作用且独立于Mre11核酸酶功能的机制，帮助驱动短TNRs的扩增。
