title: The Gata1 5' region harbors distinct cis-regulatory modules that direct gene activation in erythroid cells and gene inactivation in HSCs.
abstract: GATA1 is a master regulator of hematopoietic differentiation, but Gata1 expression is inactivated in hematopoietic stem cells (HSCs). Using a bacterial artificial chromosome containing the Gata1 gene modified with green fluorescent protein (GFP) reporter, we explored the function of the 3.7-kb Gata1 upstream region (GdC region) that harbors 3 core cis-elements: Gata1 hematopoietic enhancer, double GATA-motif, and CACCC-motif. Transgenic GFP expression directed by the Gata1-BAC faithfully recapitulated the endogenous Gata1 expression pattern. However, deletion of the GdC-region eliminated reporter expression in all hematopoietic cells. To test whether the combination of the core cis-elements represents the regulatory function of the GdC-region, we replaced the region with a 659-bp minigene that linked the three cis-elements (MG-GFP). The GFP reporter expression directed by the MG-GFP BAC fully recapitulated the erythroid-megakaryocytic Gata1 expression. However, the GFP expression was aberrantly increased in the HSCs and was associated with decreases in DNA methylation and abundant GATA2 binding to the transgenic MG-GFP allele. The 3.2-kb sequences interspaced between the Gata1 hematopoietic enhancer and the double GATA-motif were able to recruit DNA methyltransferase 1, thereby exerting a cis-repressive function in the HSC-like cell line. These results indicate that the 3.2-kb interspacing sequences inactivate Gata1 by maintaining DNA-methylation in the HSCs.
authors.affiliation: Department of Medical Biochemistry and.
authors.forename: Jun
authors.initials: J
authors.lastname: Takai
authors.forename: Takashi
authors.initials: T
authors.lastname: Moriguchi
authors.forename: Mikiko
authors.initials: M
authors.lastname: Suzuki
authors.forename: Lei
authors.initials: L
authors.lastname: Yu
authors.forename: Kinuko
authors.initials: K
authors.lastname: Ohneda
authors.forename: Masayuki
authors.initials: M
authors.lastname: Yamamoto
mesh_terms: D000818:Animals; D019070:Cell Lineage; D002478:Cells, Cultured / Q000378:metabolism; D022202:Chromosomes, Artificial, Bacterial; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism; D019175:DNA Methylation / Q000235:genetics; D004742:Enhancer Elements, Genetic / Q000235:genetics; D041905:Erythroid Cells / Q000378:metabolism*; D004920:Erythropoiesis / Q000235:genetics; D050982:GATA1 Transcription Factor / Q000235:genetics* / Q000502:physiology; D050989:GATA2 Transcription Factor / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000235:genetics*; D020868:Gene Silencing; D017930:Genes, Reporter; D005813:Genes, Synthetic; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000196:embryology; D008533:Megakaryocytes / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D059372:Nucleotide Motifs / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D017384:Sequence Deletion; D015533:Transcriptional Activation / Q000235:genetics
chemical_list: D050982:GATA1 Transcription Factor; D050989:GATA2 Transcription Factor; C494689:Gata1 protein, mouse; C494712:Gata2 protein, mouse; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases