Gfi1 expression is controlled by five distinct regulatory regions spread over 100 kilobases, with Scl/Tal1, Gata2, PU.1, Erg, Meis1, and Runx1 acting as upstream regulators in early hematopoietic cells.
The growth factor independence 1 (Gfi1) gene was originally discovered in the hematopoietic system, where it functions as a key regulator of stem cell homeostasis, as well as neutrophil and T-cell development. Outside the blood system, Gfi1 is essential for inner-ear hair and intestinal secretory cell differentiation. To understand the regulatory hierarchies within which Gfi1 operates to control these diverse biological functions, we used a combination of comparative genomics, locus-wide chromatin immunoprecipitation assays, functional validation in cell lines, and extensive transgenic mouse assays to identify and characterize the complete ensemble of Gfi1 regulatory elements. This concerted effort identified five distinct regulatory elements spread over 100kb each driving expression in transgenic mice to a subdomain of endogenous Gfi1. Detailed characterization of an enhancer 35 kb upstream of Gfi1 demonstrated activity in the dorsal aorta region and fetal liver in transgenic mice, which was bound by key stem cell transcription factors Scl/Tal1, PU.1/Sfpi1, Runx1, Erg, Meis1, and Gata2. Taken together, our results reveal the regulatory regions responsible for Gfi1 expression and importantly establish that Gfi1 expression at the sites of hematopoietic stem cell (HSC) emergence is controlled by key HSC regulators, thus integrating Gfi1 into the wider HSC regulatory networks.
University of Cambridge, Department of Haematology, Cambridge Institute for Medical Research, Cambridge, United Kingdom.
Nicola K
NK
Wilson
Richard T
RT
Timms
Sarah J
SJ
Kinston
Yi-Han
YH
Cheng
S Helen
SH
Oram
Josette-Renee
JR
Landry
Joanne
J
Mullender
Katrin
K
Ottersbach
Berthold
B
Gottgens
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D006410:Hematopoiesis / Q000235:genetics*; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D006413:Hematopoietic System / Q000378:metabolism; D018398:Homeodomain Proteins; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D012045:Regulatory Sequences, Nucleic Acid; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D018398:Homeodomain Proteins; D036341:Intercellular Signaling Peptides and Proteins; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; C000616391:MEIS1 protein, human; C000616392:Meis1 protein, mouse; C067143:TAL1 protein, human