Microsatellite instability (MSI) at simple repeated sequences characterises a distinct mechanism of carcinogenesis in hereditary nonpolyposis colorectal cancer (HNPCC), as well as sporadic colorectal cancers displaying MSI. Such MSI is associated with mutations of hMSH2 and hMLH1, and somatic frameshift mutations in TGF-beta RII, IGFIIR, BAX, hMSH3 and hMSH6 at simple repeated sequences in coding regions. The aim of this study was to look for a correlation between mutations in mismatch repair genes and frameshift mutations in colorectal cancer cell lines with MSI. Using 22 colorectal cancer cell lines, we examined the MSI status at mononucleotide repeat microsatellite markers and mutations in hMSH2 and hMLH1, TGF-beta RII, IGFIIR, BAX, hMSH3 and hMSH6. Thirteen of 22 lines (59%) displayed MSI. In these 13 lines showing MSI, 10 lines (77%) had mutations in TGF-beta RII, nine lines (69%) in BAX, seven lines (54%) in hMSH6, and six lines (46%) in hMSH3, while mutations in the IGFIIR gene were identified in only two lines (15%). Of the 13 lines with MSI, six lines (46%) harboured mutations/deletions in hMSH2 (two nonsense mutations, three deletions and one no expression of transcripts) and three of these cell lines (50%) had mutations both in the hMSH2 and hMSH3 genes. Two cell lines (15%) had mutations/deletions in hMLH1 (one missense mutation and one deletion) and these two cell lines also had mutations in hMSH3. One line had a mutation in hMSH3 only, although this line showed MSI and had mutations in TGF-beta RII, IGFIIR and BAX. All lines with mutations in hMLH1, hMSH2, TGF-beta RII, IGFIIR, BAX and hMSH3 genes showed MSI. However, of the nine lines without MSI, two (22%) had homozygous mutations in hMSH6. In these two cell lines, no mutations were identified in hMLH1, hMSH2, TGF-beta RII, IGFIIR, BAX and hMSH3. Our results indicate that mutations in hMLH1, hMSH2 and hMSH3 are associated with MSI, but mutations in hMSH6 are not. We conclude that mutations in hMSH6 alone are not sufficient to cause MSI, although protein functional effects of these mutations should still be examined.