Shimodaira H, Filosi N, Shibata H, Suzuki T, Radice P, Kanamaru R, Friend S H, Kolodner R D, Ishioka C
Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan.
Nat Genet. 1998 Aug;19(4):384-9. doi: 10.1038/1277.
Hereditary non-polyposis colorectal cancer (HNPCC; OMIM 120435-6) is a cancer-susceptibility syndrome linked to inherited defects in human mismatch repair (MMR) genes. Germline missense human MLH1 (hMLH1) mutations are frequently detected in HNPCC (ref. 3), making functional characterization of mutations in hMLH1 critical to the development of genetic testing for HNPCC. Here, we describe a new method for detecting mutations in hMLH1 using a dominant mutator effect of hMLH1 cDNA expressed in Saccharomyces cerevisiae. The majority of hMLH1 missense mutations identified in HNPCC patients abolish the dominant mutator effect. Furthermore, PCR amplification of hMLH1 cDNA from mRNA from a HNPCC patient, followed by in vivo recombination into a gap expression vector, allowed detection of a heterozygous loss-of-function missense mutation in hMLH1 using this method. This functional assay offers a simple method for detecting and evaluating pathogenic mutations in hMLH1.
遗传性非息肉病性结直肠癌（HNPCC；OMIM 120435 - 6）是一种癌症易感性综合征，与人类错配修复（MMR）基因的遗传缺陷有关。在HNPCC中经常检测到种系错义人类MLH1（hMLH1）突变（参考文献3），因此对hMLH1突变进行功能表征对于HNPCC基因检测的发展至关重要。在此，我们描述了一种利用在酿酒酵母中表达的hMLH1 cDNA的显性诱变效应来检测hMLH1突变的新方法。在HNPCC患者中鉴定出的大多数hMLH1错义突变消除了显性诱变效应。此外，从HNPCC患者的mRNA中进行hMLH1 cDNA的PCR扩增，然后在体内重组到缺口表达载体中，使用该方法可检测到hMLH1中的杂合功能丧失错义突变。这种功能测定法为检测和评估hMLH1中的致病突变提供了一种简单方法。
