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R.K. Hyde, A.E. Griep; Specific and Dynamic Association of E2F Family Members with Target Gene Promoters in the Mouse Ocular Lens . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4536.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Cell cycle regulation, controlled in part by the activity of the retinoblastoma protein (pRB), is essential for maintaining the normal patterns of growth and differentiation of lens. It is thought that the expression of many genes involved in cell cycle progression are regulated by complexes containing a member of the pRB family and a member of the E2F family of transcription factors. Studies in the lens support this model. To date it is not known if individual E2F target genes are regulated by specific pRB:E2F combinations and if the regulation of a single promoter is fixed, or can change under different conditions. Methods: To characterize the profile of E2F family members associated with the promoters of various E2F target genes in the context of the normal mouse lens, we performed chromatin immunoprecipitation (ChIP) experiments. Chromatin was isolated from the lenses of neonatal or adult mice, and E2F–DNA complexes immunoprecipitated with antibodies specific for E2F1, 2, 3a, 4, 5, or 6. Promoters for various E2F target genes were PCR amplified. Products were analyzed by Southern blot and quantified by phosphoimager. Results: In lenses of nontransgenic neonatal mice, different promoters show different profiles of associated E2F family members. p19ARF and cdc2 are most strongly associated with E2F3a, while n–myc shows nearly equal association with both E2F3a and E2F5, and bmyb with E2F2, 3a and 4. In lenses of adult mice, these profiles appear to be different. The pattern of associated E2Fs changes in the lenses of E2F1–null neonatal mice as promoters show varying increases in associated E2F2, and/or E2F5.Conclusions: In the lens, different profiles of E2Fs are associated with the p19ARF, bmyb, cdc2 and n–myc promoters; however, with age or the loss of E2F1, the patterns change. These data suggest that specific combinations of E2Fs preferentially regulate individual E2F target genes and that the patterns can be differentially altered. This implies that E2F regulation of individual promoters is both specific and dynamic, allowing for the tightly controlled target gene expression required for lens development and maturation.
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