Functions of MSG1 Family Transcription Activators

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DESCRIPTION (provided by applicant): Members of the CITED family, formerly called the MSG 1 family, function as transcriptional coregulators. By interacting with both sequence-specific transcription factors and CBP/p300, the CITED proteins affect CBP/p300-dependent transcription. Importantly, the coregulating activity of the CITED proteins show remarkable gene-specificity; binding of a CITED-interacting transcription factor to a promoter is necessary - but not sufficient - for recruitment of the CITED proteins to promoters. Since only three genes - TGF-a, c-MYC, and glycoprotein a-subunit gene - have been identified as CITED-regulated genes so far, our knowledge of biological roles and molecular mechanisms of functions of the CITED proteins is still very limited. Therefore, in this project, attempts will be made to identify more CITED-regulated genes and to elucidate biochemical basis of the gene-specific coregulating activity of the CITED proteins. In Specific Aim 1.1, to obtain insights into CITED-regulated genes, phenotypes of knockout mice lacking the cited genes (cited1, cited2, and cited4) will be examined. To identify candidate CITED-regulated genes, mRNA expression profiles of the affected tissues will be determined. In Specific Aim 1.2, to obtain clues for identifying novel CITED-regulated promoters, attempts will be made to isolate genomic DNA fragments that recruit the CITED proteins by chromatin immunoprecipitation. In Specific Aim 1.3, roles of CITED4 in transcription of three candidate CITED4-regulated genes will be examined. Amounts of the mRNA transcripts of those candidates increased along with induced overexpression of CITED4 in human osteosarcoma cells. Attempts will be made to demonstrate direct involvement of CITED4 in transcriptional activation of these genes by cell culture-based analyses: reporter assays and chromatin immunoprecipitation. Physiological roles of CITED4 in their regulation will be evaluated using cited4 knockout mice. Finally, in Specific Aim 2, recruitment and modifications of protein factors to known CITED 1-regulated promoters in cultured cells will be examined by chromatin immunoprecipitation.