title: Identification of KAP-1-associated complexes negatively regulating the Ey and β-major globin genes in the β-globin locus.
abstract: Deregulations of erythroid differentiation may lead to erythroleukemia and other hemoglobinopathies, yet the molecular mechanisms underlying these events are not fully understood. Here, we found that KAP-1-associated complexes contribute to the regulation of the β-globin locus, the key events of erythroid differentiation. We show that RNAi-mediated knockdown of KAP-1 in mouse erythroleukemia (MEL) cells increases expression of the Ey and β-major globin genes during hexamethylenebisacetamide (HMBA) induced differentiation process. This indicates that at least part of KAP-1-associated complexes negatively regulates β-globin gene expression during definitive erythroid differentiation. ChIP-PCR analysis revealed that one or more KAP-1-associated complexes are targeted to the promoter region of the Ey and beta-major globin genes. Since KAP-1 is only a scaffold molecule, there must be some transcriptional regulators allowing its targeted recruitment to the β-globin locus. To further discover these novel regulators, proteins interacting with KAP-1 were isolated by endogenous immunoprecipitation and identified by LC-ESI-MS/MS. Among the proteins identified, MafK and Zfp445 were studied further. We found that KAP-1 may contribute to the repression of Ey and β-major globin gene transcription through recruitment to the promoters of these two genes, mediated by the interaction of KAP-1 with either Zfp445 or MafK, respectively.
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China.
authors.forename: Dong
authors.initials: D
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China; College of Life Science and Bio-engineering, Beijing University of Technology, Beijing 100022, PR China.
authors.forename: Zhanfu
authors.initials: Z
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China.
authors.forename: Weiran
authors.initials: W
authors.lastname: Lin
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China.
authors.forename: Juntao
authors.initials: J
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China.
authors.forename: Chunyan
authors.initials: C
authors.lastname: Tian
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China.
authors.forename: Handong
authors.initials: H
authors.lastname: Wei
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China. Electronic address: jiangying304@hotmail.com.
authors.forename: Ying
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, PR China; Institutes of Biomedical Sciences and Department of Chemistry, Fudan University, Shanghai 200032, PR China. Electronic address: hefc@nic.bmi.ac.cn.
authors.forename: Fuchu
authors.initials: F
authors.lastname: He
mesh_terms: D000081:Acetamides / Q000737:chemistry; D000818:Animals; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D002853:Chromatography, Liquid; D004268:DNA-Binding Proteins / Q000378:metabolism; D004912:Erythrocytes / Q000166:cytology; D005136:Eye Proteins / Q000378:metabolism*; D020869:Gene Expression Profiling; D015973:Gene Expression Regulation, Leukemic; D005982:Glutathione Transferase / Q000378:metabolism; D057809:HEK293 Cells; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D050985:MafK Transcription Factor / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D053719:Tandem Mass Spectrometry; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D000076183:Tripartite Motif-Containing Protein 28; D016335:Zinc Fingers; D055544:beta-Globins / Q000378:metabolism*
chemical_list: D000081:Acetamides; D004268:DNA-Binding Proteins; D005136:Eye Proteins; D018398:Homeodomain Proteins; D050985:MafK Transcription Factor; C494696:Mafk protein, mouse; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; C105441:Rax protein, mouse; D012097:Repressor Proteins; D014157:Transcription Factors; C000599170:Zfp445 protein, mouse; D055544:beta-Globins; C103190:Trim28 protein, mouse; D000076183:Tripartite Motif-Containing Protein 28; D005982:Glutathione Transferase; C014026:hexamethylene bisacetamide