Characterization of megakaryocyte GATA1-interacting proteins: the corepressor ETO2 and GATA1 interact to regulate terminal megakaryocyte maturation.
The transcription factor GATA1 coordinates timely activation and repression of megakaryocyte gene expression. Loss of GATA1 function results in excessive megakaryocyte proliferation and disordered terminal platelet maturation, leading to thrombocytopenia and leukemia in patients. The mechanisms by which GATA1 does this are unclear. We have used in vivo biotinylated GATA1 to isolate megakaryocyte GATA1-partner proteins. Here, several independent approaches show that GATA1 interacts with several proteins in the megakaryocyte cell line L8057 and in primary megakaryocytes. They include FOG1, the NURD complex, the pentameric complex containing SCL/TAL-1, the zinc-finger regulators GFI1B and ZFP143, and the corepressor ETO2. Knockdown of ETO2 expression promotes megakaryocyte differentiation and enhances expression of select genes expressed in terminal megakaryocyte maturation, eg, platelet factor 4 (Pf4). ETO2-dependent direct repression of the Pf4 proximal promoter is mediated by GATA-binding sites and an E-Box motif. Consistent with this, endogenous ETO2, GATA1, and the SCL pentameric complex all specifically bind the promoter in vivo. Finally, as ETO2 expression is restricted to immature megakaryocytes, these data suggest that ETO2 directly represses inappropriate early expression of a subset of terminally expressed megakaryocyte genes by binding to GATA1 and SCL.
Hamlett
Isla
I
Medical Research Council (MRC) Molecular Haematology Unit and Department of Haematology, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, United Kingdom.
Draper
Julia
J
Strouboulis
John
J
Iborra
Francisco
F
Porcher
Catherine
C
Vyas
Paresh
P
D000818:Animals; D001483:Base Sequence; D019921:Biotinylation; D002454:Cell Differentiation; D002460:Cell Line; D024721:E-Box Elements / Q000235:genetics; D050982:GATA1 Transcription Factor / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D008533:Megakaryocytes / Q000166:cytology* / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000302:isolation & purification / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010978:Platelet Factor 4 / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D021381:Protein Transport; D012097:Repressor Proteins / Q000378:metabolism*; D015203:Reproducibility of Results; D019809:Streptavidin / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation / Q000235:genetics
C117411:Cbfa2t3 protein, mouse; D050982:GATA1 Transcription Factor; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D010978:Platelet Factor 4; D019809:Streptavidin