A genome-wide screen identifies <i>Runx2</i> as a novel regulator of hematopoietic stem cell expansion and T-cell commitment
Abstract
Abstract Self-renewing multipotent hematopoietic stem cells (HSCs) are a rare but important cell population that can reconstitute the entire blood and immune system after transplantation. Due to their rarity, it has been difficult to comprehensively study the mechanisms regulating HSC activity. However, recent improvements in hematopoietic stem and progenitor cell (HSPC) culture methods using polyvinyl alcohol–based media now facilitate large-scale ex vivo HSC expansion. Here, we performed a genome-wide CRISPR knockout (KO) screen in primary mouse HSPCs to discover novel regulators of ex vivo expansion. The screen identified Runx2 as a strong negative regulator of HSC expansion, which we validated using ex vivo and in vivo assays. Loss of Runx2 increased the frequency of immunophenotypic HSCs in HSPC cultures by approximately threefold. After expansion, these Runx2-KO HSCs engrafted at approximately fivefold higher levels in transplantation assays. Noncultured Runx2-KO HSCs also displayed enhanced reconstitution potential, but loss of Runx2 did not alter blood parameters. Notably, however, T-cell reconstitution was diminished from Runx2-KO HSCs, and we further validated an additional role for Runx2 in T-cell commitment using ex vivo and in vivo assays. In summary, we have identified a multifaceted role for Runx2 in HSCs, as a negative regulator of HSC self-renewal and as a facilitator of T-cell commitment. These results contribute to our understanding of the transcriptional regulation of hematopoiesis and HSC therapies.
Article Details
Authors (17)
Grace A. Meaker
Matthew Nicholls
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Catherine Chahrour
1Medical Research Council Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Ian Hsu
Alastair Smith
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Yavor Bozhilov
Maurice Leung
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Hugo Vassort
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Leonid Olender
Oliver Beaven
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Xinran Huang
Elizabeth J. Brown
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Marlies Vanden Bempt
1Medical Research Council (MRC) Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, United Kingdom
Hwei Minn Khoo
Joydeep Bhadury
Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine
Thomas A. Milne
MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine
Adam C. Wilkinson