CLONING, STRUCTURAL CHARACTERIZATION, AND CHROMOSOMAL LOCALIZATION OFTHE HUMAN ORTHOLOGUE OF SACCHAROMYCES-CEREVISIAE MSH5 GENE

We have cloned and characterized the human orthologue of the Saccharom yces cerevisiae MutS homologue 5 (MSH5) cDNA, as well as the human gen e that encodes the MSH5 cDNA, as a step toward understanding the molec ular genetic mechanisms involved in the biological function of this no vel human protein. The identified cDNA contains a 2505-bp opera readin g frame (ORF) that encodes an 834-amino-acid polypeptide with a predic ted molecular mass of 92.9 kDa. The amino acid sequence encoded by thi s cDNA includes sequence motifs that are conserved in all known MutS h omologues existing in bacteria to humans. The cDNA appears, on the bas is of amino acid sequence analysis, to be a member of the MutS family and shares 30% sequence identity with that of S. cerevisiae MSR5, a ye ast gene that plays a critical role in facilitating crossover during m eiosis. Northern blot analysis demonstrated the presence of a 2.9-kb h uman MSH5 mRNA species in all human tissues tested, but the highest ex pression was in human testis, an organ containing cells that undergo c onstant DNA synthesis and meiosis. The expression pattern of human MSH 5 resembled that of the previously identified human MutS homologues MS H2, MSH3, and MSH6-genes that are involved in the pathogenesis of here ditary nonpolyposis colorectal cancer (HNPCC). In an effort to expedit e the search for potential disease association with this new human Mut S homologue, we have also determined the chromosomal location and stru cture of the human MSH5 Focus. Sequence and structural characterizatio n demonstrated that MSH5 spans approximately 25 kb and contains 26 exo ns that range in length from 36 bp for exon 8 to 254 bp for exon 25. M SH5 has been mapped to human chromosome band 6p21.3 by fluorescence in situ hybridization. Knowledge of the sequence and gene structure of M SH5 will now enable studies of the possible roles MSH5 may play in mei osis and/or DNA replicative mismatch repair. (C) 1998 Academic Press.