Expression of the DNA mismatch repair proteins hMLH1 and hPMS2 in normal human tissues.

D Fink,Sb Howell,S Nebel,H Zheng,Hk Kim,Rd Christen,S Aebi

doi:10.1038/bjc.1997.480

Abstract

hMLH1 and hPMS2 are part of the DNA mismatch repair complex. Mutations in these genes have been linked to hereditary non-polyposis colon cancer; they also occur in a variety of sporadic cancers. Western blot analysis and immunohistochemistry demonstrated that hMLH1 and hPMS2 are widely expressed nuclear proteins with a distribution pattern very similar to that previously described for hMSH2. These observations showing similar localization of hMLH1 and hPMS2 with hMSH2 are consistent with the biochemical function of these proteins in DNA mismatch repair.

Highlights

Microsatellite instability has been observed in most tumours arising in patients with hereditary non-polyposis colorectal cancer (HNPCC) and in many sporadic colon, gastric, endometrial, ovarian and small-cell lung carcinomas
MSH2 and GTBP form a heterodimer that binds to mismatched bases (Palombo et al, 1995) and that serves to recruit a heterodimer of hMLH1 and hPMS2 and free hPMSl to the complex (Prolla et al, 1994; Li and Modrich, 1995)
The PMS2 antibody detected a single protein of Mr 105 000, which is consistent with the reported molecular weight of hPMS2 (Figure 1B)

Summary

Methods

Cell lines and biopsy specimensThe hMLH1-deficient human colorectal adenocarcinoma cell line HCT1 16 was obtained from the American Type CultureReceived 6 December 1996 Revised 24 February 1997 Accepted 13 March 1997Correspondence to: D FinkCollection (ATCC CCL 247); sublines complemented with chromosome 3 (HCTl 16+ch3) and chromosome 2 (HCTl 16+ch2) were obtained from Drs CR Boland and M Koi. The hMLH1-deficient human colorectal adenocarcinoma cell line HCT1 16 was obtained from the American Type Culture. Collection (ATCC CCL 247); sublines complemented with chromosome 3 (HCTl 16+ch3) and chromosome 2 (HCTl 16+ch2) were obtained from Drs CR Boland and M Koi. HCT1 16+ch cells lack hMLHI, whereas HCT116+ch is complemented by microcell fusion transfer of chromosome 3 and expresses wildtype hMLHl (Koi et al, 1994). The cell lines were maintained in Iscove's modified Dulbecco's medium (Irvine Scientific, Irvine, CA, USA) supplemented with 2 mM L-glutamine and 10% heatinactivated fetal bovine serum. The chromosome-complemented lines were maintained in medium supplemented with geneticine (400 jg ml-') (Life Technologies, Gaithersburg, MD, USA). Frozen tissues were obtained from surgical resections and stored at -70° C until used

Results

Discussion

Conclusion