Differentiation and injury-repair signals modulate the interaction of E2F and pRB proteins with novel target genes in keratinocytes.
E2F transcription factors are central to epidermal morphogenesis and regeneration after injury. The precise nature of E2F target genes involved in epidermal formation and repair has yet to be determined. Identification of these genes is essential to understand how E2F proteins regulate fundamental aspects of epidermal homeostasis and transformation. We have conducted a genome-wide screen using CpG island microarray analysis to identify novel promoters bound by E2F3 and E2F5 in human keratinocytes. We further characterized several of these genes, and determined that multiple E2F and retinoblastoma (pRb) family proteins associate with them in exponentially proliferating cells. We also assessed the effect on E2F and pRb binding to those genes in response to differentiation induced by bone morphogenetic protein-6 (BMP-6), or to activation of repair mechanisms induced by transforming growth factor-beta (TGF-beta). These studies demonstrate promoter- and cytokine-specific changes in binding profiles of E2F and/or pRb family proteins. For example, E2F1, 3, 4 and p107 were recruited to the N-myc promoter in cells treated with BMP-6, whereas E2F1, 3, 4, 5, p107 and p130 were bound to this promoter in the presence of TGF-beta. Functionally, these different interactions resulted in transcriptional repression by BMP-6 and TGF-beta of the N-myc gene, via mechanisms that involved E2F binding to the promoter and association with pRb-family proteins. Thus, multiple combinations of E2F and pRb family proteins may associate with and transcriptionally regulate a given target promoter in response to differentiation and injury-repair stimuli in epidermal keratinocytes.
Chang
Wing Y
WY
Department of Physiology and Pharmacology, Regulatory Biology and Functional Genomics Research Group, Siebens-Drake Research Institute, Canada.
Andrews
Joseph
J
Carter
David E
DE
Dagnino
Lina
L
D000818:Animals; D064127:Aurora Kinases; D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D004247:DNA / Q000235:genetics / Q000378:metabolism; D050699:E2F3 Transcription Factor / Q000378:metabolism*; D050700:E2F5 Transcription Factor / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism*; D050720:Retinoblastoma-Like Protein p107 / Q000378:metabolism; D050717:Retinoblastoma-Like Protein p130 / Q000378:metabolism; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053773:Transforming Growth Factor beta1
C525816:BMP6 protein, human; C525817:Bmp6 protein, mouse; D055418:Bone Morphogenetic Protein 6; D019485:Bone Morphogenetic Proteins; D050699:E2F3 Transcription Factor; C098319:E2F3 protein, human; D050700:E2F5 Transcription Factor; C092558:E2F5 protein, human; D016271:Proto-Oncogene Proteins c-myc; D016160:Retinoblastoma Protein; D050720:Retinoblastoma-Like Protein p107; D050717:Retinoblastoma-Like Protein p130; C508960:TGFB1 protein, human; C508961:Tgfb1 protein, mouse; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D004247:DNA; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases