title: Mechanism governing a stem cell-generating cis-regulatory element.
abstract: The unremitting demand to replenish differentiated cells in tissues requires efficient mechanisms to generate and regulate stem and progenitor cells. Although master regulatory transcription factors, including GATA binding protein-2 (GATA-2), have crucial roles in these mechanisms, how such factors are controlled in developmentally dynamic systems is poorly understood. Previously, we described five dispersed Gata2 locus sequences, termed the -77, -3.9, -2.8, -1.8, and +9.5 GATA switch sites, which contain evolutionarily conserved GATA motifs occupied by GATA-2 and GATA-1 in hematopoietic precursors and erythroid cells, respectively. Despite common attributes of transcriptional enhancers, targeted deletions of the -2.8, -1.8, and +9.5 sites revealed distinct and unpredictable contributions to Gata2 expression and hematopoiesis. Herein, we describe the targeted deletion of the -3.9 site and mechanistically compare the -3.9 site with other GATA switch sites. The -3.9(-/-) mice were viable and exhibited normal Gata2 expression and steady-state hematopoiesis in the embryo and adult. We established a Gata2 repression/reactivation assay, which revealed unique +9.5 site activity to mediate GATA factor-dependent chromatin structural transitions. Loss-of-function analyses provided evidence for a mechanism in which a mediator of long-range transcriptional control [LIM domain binding 1 (LDB1)] and a chromatin remodeler [Brahma related gene 1 (BRG1)] synergize through the +9.5 site, conferring expression of GATA-2, which is known to promote the genesis and survival of hematopoietic stem cells.
authors.affiliation: Department of Cell and Regenerative Biology, Wisconsin Institutes for Medical Research, Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705.
authors.forename: Rajendran
authors.initials: R
authors.lastname: Sanalkumar
authors.forename: Kirby D
authors.initials: KD
authors.lastname: Johnson
authors.forename: Xin
authors.initials: X
authors.lastname: Gao
authors.forename: Meghan E
authors.initials: ME
authors.lastname: Boyer
authors.forename: Yuan-I
authors.initials: YI
authors.lastname: Chang
authors.forename: Kyle J
authors.initials: KJ
authors.lastname: Hewitt
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Emery H
authors.initials: EH
authors.lastname: Bresnick
mesh_terms: D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics; D002478:Cells, Cultured; D004742:Enhancer Elements, Genetic; D050989:GATA2 Transcription Factor / Q000378:metabolism*; D006410:Hematopoiesis; D006801:Humans; D007438:Introns; D051379:Mice; D008969:Molecular Sequence Data; D012045:Regulatory Sequences, Nucleic Acid; D012689:Sequence Homology, Nucleic Acid; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism
chemical_list: D050989:GATA2 Transcription Factor; C494712:Gata2 protein, mouse
keywords: HSCs; cis element