Differentiation and Injury-Repair Signals Modulate the Interaction of E2F and pRB Proteins with Novel Target Genes in Keratinocytes

Abstract

E2F transcription factors are central to epidermal morphogenesis and regeneration afterinjury. The precise nature of E2F target genes involved in epidermal formation and repair has yetto be determined. Identification of these genes is essential to understand how E2F proteinsregulate fundamental aspects of epidermal homeostasis and transformation. We have conducted agenome-wide screen using CpG island microarray analysis to identify novel promoters bound byE2F3 and E2F5 in human keratinocytes. We further characterized several of these genes, anddetermined that multiple E2F and retinoblastoma (pRb) family proteins associate with them inexponentially proliferating cells. We also assessed the effect on E2F and pRb binding to thosegenes in response to differentiation induced by bone morphogenetic protein-6 (BMP-6), or toactivation of repair mechanisms induced by transforming growth factor-β (TGF-β). Thesestudies demonstrate promoter- and cytokine-specific changes in binding profiles of E2F and/orpRb family proteins. For example, E2F1, 3, 4 and p107 were recruited to the N-myc promoter incells treated with BMP-6, whereas E2F1, 3, 4, 5, p107 and p130 were bound to this promoter inthe presence of TGF-β. Functionally, these different interactions resulted in transcriptionalrepression by BMP-6 and TGF-β of the N-myc gene, via mechanisms that involved E2F bindingto the promoter and association with pRb-family proteins. Thus, multiple combinations of E2Fand pRb family proteins may associate with and transcriptionally regulate a given target promoterin response to differentiation and injury-repair stimuli in epidermal keratinocytes.