Regulation of the human MSH6 gene by the Sp1 transcription factor and alteration of promoter activity and expression by polymorphisms.
Defects in human DNA mismatch repair have been reported to underlie a variety of hereditary and sporadic cancer cases. We characterized the structure of the MSH6 promoter region to examine the mechanisms of transcriptional regulation of the MSH6 gene. The 5'-flanking region of the MSH6 gene was found to contain seven functional Sp1 transcription factor binding sites that each bind Sp1 and Sp3 and contribute to promoter activity. Transcription did not appear to require a TATA box and resulted in multiple start sites, including two major start sites and at least nine minor start sites. Three common polymorphisms were identified in the promoter region (-557 T-->G, -448 G-->A, and -159 C-->T): the latter two were always associated, and each of these functionally inactivated a different Sp1 site. The polymorphic allele -448 A -159 T was demonstrated to be a common Caucasian polymorphism found in 16% of Caucasians and resulted in a five-Sp1-site promoter that had 50% less promoter activity and was more sensitive to inactivation by DNA methylation than the more common seven Sp1 site promoter allele, which was only partially inactivated by DNA methylation. In cell lines, this five-Sp1-site polymorphism resulted in reduced MSH6 expression at both the mRNA and protein level. An additional 2% of Caucasians contained another polymorphism, -210 C-->T, which inactivated a single Sp1 site that also contributes to promoter activity.
Ludwig Institute for Cancer Research. University of California-San Diego School of Medicine, CMME 3058, 9500 Gilman Drive, La Jolla, CA 92093, USA. igazzoli@ucsd.edu
Isabella
I
Gazzoli
Richard D
RD
Kolodner
D000483:Alleles; D000818:Animals; D020137:Base Pair Mismatch; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D045744:Cell Line, Tumor; D006224:Cricetinae; D004247:DNA / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D004260:DNA Repair / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D014644:Genetic Variation; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics; D011110:Polymorphism, Genetic; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection
D004268:DNA-Binding Proteins; C094443:G-T mismatch-binding protein; D016329:Sp1 Transcription Factor; D004247:DNA