title: Enhanced detection of deleterious and other germline mutations of hMSH2 and hMLH1 in Japanese hereditary nonpolyposis colorectal cancer kindreds.
abstract: Hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal, dominantly inherited cancer-prone syndrome. Here, we describe a novel and efficient approach for screening mutations of two major HNPCC susceptibility genes, hMSH2 and hMLH1. The system consists of RNA extraction from whole blood treated with the translation inhibitor, followed by long RT-PCR of the entire coding regions combined with direct sequencing. In analysis of 15 kindreds suspicious for HNPCC, 8 samples were subjected to analysis after puromycin treatment and 7 samples were analyzed without puromycin treatment. Three deleterious mutations were detected in the kindreds with puromycin treatment, while none were observed in those without puromycin. Signals from mutated alleles were enhanced after puromycin treatment and easily distinguished from the wild-type allele, achieved by suppression of nonsense-mediated mRNA decay. Furthermore, 12 other mutations were detected in 15 kindreds. The system is considered to be a reliable and useful approach for detecting germline mutations of hMSH2 and hMLH1 in HNPCC kindreds.
authors.affiliation: Division of Clinical Laboratory, National Cancer Center Hospital, Tokyo, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Nomura
authors.forename: K
authors.initials: K
authors.lastname: Sugano
authors.forename: H
authors.initials: H
authors.lastname: Kashiwabara
authors.forename: T
authors.initials: T
authors.lastname: Taniguchi
authors.forename: N
authors.initials: N
authors.lastname: Fukayama
authors.forename: S
authors.initials: S
authors.lastname: Fujita
authors.forename: T
authors.initials: T
authors.lastname: Akasu
authors.forename: Y
authors.initials: Y
authors.lastname: Moriya
authors.forename: S
authors.initials: S
authors.lastname: Ohhigashi
authors.forename: T
authors.initials: T
authors.lastname: Kakizoe
authors.forename: T
authors.initials: T
authors.lastname: Sekiya
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000483:Alleles; D000841:Anisomycin / Q000494:pharmacology; D002352:Carrier Proteins; D003123:Colorectal Neoplasms, Hereditary Nonpolyposis / Q000235:genetics*; D003513:Cycloheximide / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D005091:Exons; D005192:Family Health; D016368:Frameshift Mutation; D017353:Gene Deletion; D005787:Gene Frequency; D018095:Germ-Line Mutation; D006801:Humans; D007564:Japan; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein; D009363:Neoplasm Proteins / Q000235:genetics*; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D011691:Puromycin / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C501486:MLH1 protein, human; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D011500:Protein Synthesis Inhibitors; D011518:Proto-Oncogene Proteins; D011691:Puromycin; D000841:Anisomycin; D003513:Cycloheximide; C497172:MSH2 protein, human; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein