Germline and somatic mutations in hMSH6 and hMSH3 in gastrointestinal cancers of the microsatellite mutator phenotype

Hereditary and sporadic gastrointestinal cancer of the microsatellite mutat or phenotype (MMP) is characterized by a remarkable genomic instability at simple repeated sequences. The genomic instability is often caused by germl ine and somatic mutations in DNA mismatch repair (MMR) genes hMSH2 and hMLH 1. The MMP can be also caused by epigenetic inactivation of hMLH1. The MMP generates many somatic frameshift mutations in genes containing mononucleot ide repeats. We previously reported that in MMP tumors the hMSH6 and hMSH3 MMR genes often carry frameshift mutations in their (C)(8) and (A)(8) track s. respectively. We proposed that these 'secondary mutator mutations' contr ibute to a gradual manifestation of the MMP. Here we report the detection o f other frameshift. nonsense. and missense mutations in these genes in colo n and gastric cancers of the MMP. A germline frameshift mutation was found in hMSH6 in a colon tumor harboring another somatic frameshift mutation. Se veral germline sequence variants and somatic missense mutations at conserve d residues were detected in hMSH6 and only one was detected in hMSH3. Of th e three hMSH6 germline variants in conserved residues, one coexisted with a somatic mutation at the (C)(8) track and another had a somatic missense mu tation. We suggest that some of these germline and somatic missense variant s are pathogenic. While biallelic hMSH6 and hMSH3 frameshift mutations were found in some tumors, many tumors seemed to contain only monoallelic mutat ions. In some tumors, these somatic monoallelic frameshift mutations at the (C)(8) and (A),., tracks were found to coexist with other somatic mutation s in the other allele, supporting their functionality during tumorigenesis. However, the low incidence of these additional somatic mutations in hMSH6 and hMSH3 leaves many tumors with only monoallelic mutations. The impact of the frameshift mutations in gene expression was studied by comparative ana lysis of RNA and protein expression in different tumor cell clones with dif ferent genotypes. The results show that the hMSH6 (C)(8) frameshift mutatio n abolishes protein expression. ruling out a dominant negative effect by a truncated protein. We suggest the functionality of these secondary monoalle lic mutator mutations in the context of an accumulative haploinsufficiency model. (C) 2001 Elsevier Science B.V. All rights reserved.