ID2 and HIF-1α collaborate to protect quiescent hematopoietic stem cells from activation, differentiation, and exhaustion
Brad L. Jakubison, Tanmoy Sarkar, Kristbjorn O. Gudmundsson, Shweta Singh, Lei Sun, Holly M. Morris, Kimberly D. Klarmann, Jonathan R. Keller
Brad L. Jakubison, Tanmoy Sarkar, Kristbjorn O. Gudmundsson, Shweta Singh, Lei Sun, Holly M. Morris, Kimberly D. Klarmann, Jonathan R. Keller
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Research Article Hematology

ID2 and HIF-1α collaborate to protect quiescent hematopoietic stem cells from activation, differentiation, and exhaustion

Abstract

Defining mechanism(s) that maintain tissue stem quiescence is important for improving tissue regeneration, cell therapies, aging, and cancer. We report here that genetic ablation of Id2 in adult hematopoietic stem cells (HSCs) promotes increased HSC activation and differentiation, which results in HSC exhaustion and bone marrow failure over time. Id2Δ/Δ HSCs showed increased cycling, ROS production, mitochondrial activation, ATP production, and DNA damage compared with Id2+/+ HSCs, supporting the conclusion that Id2Δ/Δ HSCs are less quiescent. Mechanistically, HIF-1α expression was decreased in Id2Δ/Δ HSCs, and stabilization of HIF-1α in Id2Δ/Δ HSCs restored HSC quiescence and rescued HSC exhaustion. Inhibitor of DNA binding 2 (ID2) promoted HIF-1α expression by binding to the von Hippel-Lindau (VHL) protein and interfering with proteasomal degradation of HIF-1α. HIF-1α promoted Id2 expression and enforced a positive feedback loop between ID2 and HIF-1α to maintain HSC quiescence. Thus, sustained ID2 expression could protect HSCs during stress and improve HSC expansion for gene editing and cell therapies.

Authors

Brad L. Jakubison, Tanmoy Sarkar, Kristbjorn O. Gudmundsson, Shweta Singh, Lei Sun, Holly M. Morris, Kimberly D. Klarmann, Jonathan R. Keller

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Figure 6

Loss of Id2 results in decreased expression of HIF-1α and glycolytic target genes and hypoxic status in HSCs.

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Loss of Id2 results in decreased expression of HIF-1α and glycolytic tar...
(A) Analysis of HIF-1α expression in HSCs, neutrophils, and B cells by flow cytometry and the percentage of HIF-1α expression in HSPCs. (B) Flow cytometric analysis of HIF-1α expression in Id2+/+ and Id2Δ/Δ HSCs from chimeric mice, 4 and 8 weeks after Id2 deletion. (C) Procedure to measure HIF-1α expression and hypoxic status (PIMO staining) of Id2+/+ and Id2Δ/Δ HSCs. (D) FACS-sorted Id2+/+ and Id2Δ/Δ HSCs were stained with antibodies that detect HIF-1α and imaged by confocal microscopy. (E) Pimonidazole levels in Id2+/+ and Id2Δ/Δ HSCs determined by flow cytometry. Original magnification, 40×. (F) Analysis of HIF-1α targets in Id2+/+ and Id2Δ/Δ HSCs, including genes related to glycolysis (Pdk1, Hk3), oxidative phosphorylation (OXPHOS) (Pdh1a2), and ROS (Gss). (G) Estimation of ATP levels and oxidized DNA in Id2+/+ and Id2Δ/Δ HSCs. (H) HIF-1α expression and ROS levels in ID2hi/loeYFPhi/lo HSCs. In B, E, F, G, and H, data are presented as the mean ± SEM. Comparisons between the mean values of 2 groups were evaluated using an unpaired, 1-tailed Student’s t test. *P ≤ 0.05, **P ≤ 0.01, and ***P ≤ 0.001.