title: RBF binding to both canonical E2F targets and noncanonical targets depends on functional dE2F/dDP complexes.
abstract: The retinoblastoma (RB) family of proteins regulate transcription. These proteins lack intrinsic DNA-binding activity but are recruited to specific genomic locations through interactions with sequence-specific DNA-binding factors.  The best-known target of RB protein (pRB) is the E2F transcription factor; however, many other chromatin-associated proteins have been described that may allow RB family members to act at additional sites. To gain a perspective on the scale of E2F-dependent and E2F-independent functions, we generated genome-wide binding profiles of RBF1 and dE2F proteins in Drosophila larvae. RBF1 and dE2F2 associate with a large number of binding sites at genes with diverse biological functions. In contrast, dE2F1 was detected at a smaller set of promoters, suggesting that it overrides repression by RBF1/dE2F2 at a specific subset of targets. Approximately 15% of RBF1-bound regions lacked consensus E2F-binding motifs. To test whether RBF1 action at these sites is E2F independent, we examined dDP mutant larvae that lack any functional dE2F/dDP heterodimers. As measured by chromatin immunoprecipitation-microarray analysis (ChIP-chip), ChIP-quantitative PCR (qPCR), and cell fractionation, the stable association of RBF1 with chromatin was eliminated in dDP mutants. This requirement for dDP was seen at classic E2F-regulated promoters and at promoters that lacked canonical E2F-binding sites. These results suggest that E2F/DP complexes are essential for all genomic targeting of RBF1.
authors.affiliation: Massachusetts General Hospital Cancer Center and Harvard Medical School, Charlestown, Massachusetts, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Korenjak
authors.forename: Endre
authors.initials: E
authors.lastname: Anderssen
authors.forename: Sridhar
authors.initials: S
authors.lastname: Ramaswamy
authors.forename: Johnathan R
authors.initials: JR
authors.lastname: Whetstine
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Dyson
mesh_terms: D000818:Animals; D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D029721:Drosophila Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D050684:E2F Transcription Factors / Q000235:genetics / Q000378:metabolism*; D007814:Larva / Q000378:metabolism; D009154:Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016160:Retinoblastoma Protein; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; C088053:Dp transcription factor, Drosophila; D029721:Drosophila Proteins; D050684:E2F Transcription Factors; C496298:E2f1 protein, Drosophila; C099965:Rbf protein, Drosophila; D016160:Retinoblastoma Protein; D015534:Trans-Activators; D014157:Transcription Factors