title: PIAS1 regulates CP2c localization and active promoter complex formation in erythroid cell-specific alpha-globin expression.
abstract: Data presented here extends our previous observations on α-globin transcriptional regulation by the CP2 and PIAS1 proteins. Using RNAi knockdown, we have now shown that CP2b, CP2c and PIAS1 are each necessary for synergistic activation of endogenous α-globin gene expression in differentiating MEL cells. In this system, truncated PIAS1 mutants lacking the ring finger domain recruited CP2c to the nucleus, as did wild-type PIAS1, demonstrating that this is a sumoylation-independent process. In vitro, recombinant CP2c, CP2b and PIAS1 bound DNA as a stable CBP (CP2c/CP2b/PIAS1) complex. Following PIAS1 knockdown in MEL cells, however, the association of endogenous CP2c and CP2b with the α-globin promoter simultaneously decreased. By mapping the CP2b- and CP2c-binding domains on PIAS1, and the PIAS1-binding domains on CP2b and CP2c, we found that two regions of PIAS1 that interact with CP2c/CP2b are required for its co-activator function. We propose that CP2c, CP2b, and PIAS1 form a hexametric complex with two units each of CP2c, CP2b, and PIAS1, in which PIAS1 serves as a clamp between two CP2 proteins, while CP2c binds directly to the target DNA and CP2b mediates strong transactivation.
authors.affiliation: Department of Life Science and Research Institute for Natural Sciences, College of Natural Sciences, Hanyang University, Seoul, 133-791, Korea.
authors.forename: Ho Chul
authors.initials: HC
authors.lastname: Kang
authors.forename: Ji Hyung
authors.initials: JH
authors.lastname: Chae
authors.forename: Jinseon
authors.initials: J
authors.lastname: Jeon
authors.forename: Won
authors.initials: W
authors.lastname: Kim
authors.forename: Dae Hyun
authors.initials: DH
authors.lastname: Ha
authors.forename: June Ho
authors.initials: JH
authors.lastname: Shin
authors.forename: Chan Gil
authors.initials: CG
authors.lastname: Kim
authors.forename: Chul Geun
authors.initials: CG
authors.lastname: Kim
mesh_terms: D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D041905:Erythroid Cells / Q000378:metabolism*; D006801:Humans; D051379:Mice; D011401:Promoter Regions, Genetic; D050828:Protein Inhibitors of Activated STAT / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D034622:RNA Interference; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D015533:Transcriptional Activation; D055542:alpha-Globins / Q000235:genetics*
chemical_list: D004268:DNA-Binding Proteins; D050828:Protein Inhibitors of Activated STAT; C501905:Tfcp2 protein, mouse; D014157:Transcription Factors; D055542:alpha-Globins