Lnk controls mouse hematopoietic stem cell self-renewal and quiescence through direct interactions with JAK2
Alexey Bersenev, … , Joanna Balcerek, Wei Tong
Alexey Bersenev, … , Joanna Balcerek, Wei Tong
Published July 10, 2008
Citation Information: J Clin Invest. 2008;118(8):2832-2844. https://doi.org/10.1172/JCI35808.
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Research Article Hematology

Lnk controls mouse hematopoietic stem cell self-renewal and quiescence through direct interactions with JAK2

Abstract

In addition to its role in megakaryocyte production, signaling initiated by thrombopoietin (TPO) activation of its receptor, myeloproliferative leukemia virus protooncogene (c-Mpl, or Mpl), controls HSC homeostasis and self-renewal. Under steady-state conditions, mice lacking the inhibitory adaptor protein Lnk harbor an expanded HSC pool with enhanced self-renewal. We found that HSCs from Lnk–/– mice have an increased quiescent fraction, decelerated cell cycle kinetics, and enhanced resistance to repeat treatments with cytoablative 5-fluorouracil in vivo compared with WT HSCs. We further provide genetic evidence demonstrating that Lnk controls HSC quiescence and self-renewal, predominantly through Mpl. Consistent with this observation, Lnk–/– HSCs displayed potentiated activation of JAK2 specifically in response to TPO. Biochemical experiments revealed that Lnk directly binds to phosphorylated tyrosine residues in JAK2 following TPO stimulation. Of note, the JAK2 V617F mutant, found at high frequencies in myeloproliferative diseases, retains the ability to bind Lnk. Therefore, we identified Lnk as a physiological negative regulator of JAK2 in stem cells and TPO/Mpl/JAK2/Lnk as a major regulatory pathway in controlling stem cell self-renewal and quiescence.

Authors

Alexey Bersenev, Chao Wu, Joanna Balcerek, Wei Tong

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Figure 1

Mpl–/–Lnk–/– HSCs show decreased self-renewal ability.

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Mpl–/–Lnk–/– HSCs show decreased self-renewal ability. (A) We quant...
(A) We quantified HSC frequencies of WT, Mpl–/–, Lnk–/–, and Mpl–/–Lnk–/– mice using limiting dilution cBMT. Results are pooled from 3–5 independent experiments. (B) Top: BM cells from WT, Mpl–/–, Lnk–/–, and Mpl–/–Lnk–/– mice were mixed at 1:1 ratio with CD45.1 competitor cells and transplanted into irradiated recipient mice. Chimerisms of transplanted mice were measured 1, 4, and 8 months after transplant (mean ± SD). n = 10. Middle panel: Transplant results from WT, Mpl–/–, and Mpl–/–Lnk–/– mice when mixed at 9:1 ratio with the competitors. Bottom: Transplant results from Lnk–/– BM cells when mixed at different ratios with the competitors. n = 5. *P < 0.005, WT compared with Mpl–/–Lnk–/– at the same time points, 2-tailed unequal variance t test; #P < 0.005, compared with same strain at 4 months, 2-tailed paired t test; **P < 0.05, compared with same strain at 4 months, 2-tailed paired t test. (C and D) Primary transplanted mice were sacrificed at 8 months and total BM cells were transplanted into secondary recipients. (C) Transplant results from WT, Mpl–/–, and Mpl–/–Lnk–/– mice when mixed at 9:1 ratio and Lnk–/– mice when mixed at 1:19 ratio with the competitors. (D) Transplant results from WT, Mpl–/–, and Mpl–/–Lnk–/– mice when mixed at 3:1 ratio and Lnk–/– mice when mixed at 1:99 ratio with the competitors. Chimerisms of the primary transplanted mice (mean ± SD) are shown in the left panels and those of the secondary transplant (mean ± SEM) are shown in the right panels. **P < 0.05; †P = not significant, n = 5.