Wanat Jennifer J, Singh Nikhil, Alani Eric
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
Hum Mol Genet. 2007 Feb 15;16(4):445-52. doi: 10.1093/hmg/ddl479. Epub 2007 Jan 8.
Germline mutations in the DNA mismatch repair (MMR) gene MLH1 are associated with a large percentage of hereditary non-polyposis colorectal cancers. There are approximately 250 known human mutations in MLH1. Of these, one-third are missense variants that are often difficult to characterize with regards to pathogenicity. We analysed 28 alleles of baker's yeast MLH1 that correspond to non-truncating human mutant alleles listed in online HNPCC databases, 13 of which had not been previously studied in functional assays. Using the highly sensitive lys2::InsE-A(14) reversion rate assay, we determined the MMR proficiency conferred by each allele in the S288c strain of Saccharomyces cerevisiae. Seven alleles conferred a null phenotype for MMR and eight others showed significant MMR defects, suggesting that all 15 are likely to be pathogenic in humans. In addition, we observed a strong correlation between these results, limited results from previous functional assays and clinical data. To test whether the potential pathogenicity of certain alleles depends on the genetic background of the host, we examined the mutation rates conferred by the mlh1 alleles in a second yeast strain, SK1, which is approximately 0.7% divergent from S288c. Many alleles displayed a difference in MMR efficiency between strain backgrounds with decreasing differences as the severity of the MMR defect increased. These findings suggest that genetic background can play an important role in determining the pathogenicity of MMR alleles and may explain cases of atypical colorectal cancer inheritance.
DNA错配修复（MMR）基因MLH1中的种系突变与很大比例的遗传性非息肉病性结直肠癌相关。MLH1中大约有250种已知的人类突变。其中，三分之一是错义变体，其致病性通常难以确定。我们分析了28个面包酵母MLH1等位基因，它们对应于在线HNPCC数据库中列出的非截断型人类突变等位基因，其中13个以前未在功能试验中研究过。使用高度敏感的lys2::InsE-A(14)回复率测定法，我们确定了酿酒酵母S288c菌株中每个等位基因赋予的错配修复能力。7个等位基因赋予错配修复无效表型，另外8个显示出明显的错配修复缺陷，这表明所有15个等位基因在人类中可能具有致病性。此外，我们观察到这些结果、先前功能试验的有限结果与临床数据之间存在很强的相关性。为了测试某些等位基因的潜在致病性是否取决于宿主的遗传背景，我们检查了mlh1等位基因在第二个酵母菌株SK1中赋予的突变率，该菌株与S288c的差异约为0.7%。随着错配修复缺陷严重程度的增加，许多等位基因在菌株背景之间的错配修复效率存在差异，差异逐渐减小。这些发现表明，遗传背景在确定错配修复等位基因的致病性方面可能起重要作用，并可能解释非典型结直肠癌遗传的病例。
