CLASP2 safeguards hematopoietic stem cell properties during mouse and fish development
Klaus, Anna; Clapes, Thomas; Yvernogeau, Laurent; Basu, Sreya; Weijts, Bart; Maas, Joris; Smal, Ihor; Galjart, Niels; Robin, Catherine
(2022) Cell Reports, volume 39, issue 11, pp. 1 - 28
(Article)
Abstract
Hematopoietic stem cells (HSCs) express a large variety of cell surface receptors that are associated with acquisition of self-renewal and multipotent properties. Correct expression of these receptors depends on a delicate balance between cell surface trafficking, recycling, and degradation and is controlled by the microtubule network and Golgi apparatus, whose
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roles have hardly been explored during embryonic/fetal hematopoiesis. Here we show that, in the absence of CLASP2, a microtubule-associated protein, the overall production of HSCs is reduced, and the produced HSCs fail to self-renew and maintain their stemness throughout mouse and zebrafish development. This phenotype can be attributed to decreased cell surface expression of the hematopoietic receptor c-Kit, which originates from increased lysosomal degradation in combination with a reduction in trafficking to the plasma membrane. A dysfunctional Golgi apparatus in CLASP2-deficient HSCs seems to be the underlying cause of the c-Kit expression and signaling imbalance.
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Keywords: c-Kit, CLASP2, CP: Developmental biology, embryonic aorta, Golgi integrity, hematopoietic stem cells, hemogenic endothelium, intra-aortic hematopoietic clusters, mouse, post-translational regulation, zebrafish, General Biochemistry,Genetics and Molecular Biology
ISSN: 2211-1247
Publisher: Cell Press
Note: Funding Information: The authors thank the present and past lab members for helpful discussions and technical help, especially Jean-Charles Boisset, Carla Kroon, Trung Bui, and Rutger Wielink. We also thank the Jeroen den Hertog and Jeroen Bakkers labs for providing reagents, zebrafish lines, and expertise in genome editing and the Catherine Rabouille lab for antibodies. We thank Romualdo Ciau-Uitz (Prof. Roger Patient's Laboratory) for kindly providing ISH probes. We thank Umut Akinci and Alex Maas (Department of Cell Biology, Erasmus MC) for derivation of the Clasp2−/− ESCs. We thank the Animal Facility for mouse and zebrafish care and the Optical Imaging Center for confocal microscope access (Hubrecht Institute). We thank Reinier van der Linden (Hubrecht Institute) for help with cell sorting. The authors acknowledge the use of Servier Medical Art image bank to partly create the graphical abstract. This work was supported in the C.R. and N.G. labs by a Landsteiner Stichting voor Bloedtransfusieresearch (LSBR 1025), in the N.G. lab by an NWO-TTW grant (15511), and in the C.R. lab by a European Research Council grant (ERC project number 220-H75001EU/HSCOrigin-309361), a TOP subsidy from NWO/ZonMw (912.15.017), and an UMC Utrecht “Regenerative Medicine & Stem Cells” priority research program. C.R. and N.G. conceived ideas and designed the research with help from the other authors. C.R. T.C. A.K. and L.Y. performed mouse embryo dissection. C.R. performed mouse transplantation, and T.C. analyzed the mice. T.C and A.K. performed clonogenic assays. T.C. performed LTC-IC cultures, and A.K. performed explant cultures. T.C. and A.K. performed flow cytometry analyses on mouse embryos. L.Y. performed whole-mount immunostaining and intra-aortic hematopoietic cluster counts. A.K. generated the clasp2 mutant fish lines and performed all analyses on zebrafish with help from L.Y. for flow cytometry analyses. A.K. performed mouse IF staining with guidance from L.Y. and analyzed the data with C.R. S.B. and N.G. B.W. performed all qRT-PCR on mice and cloned the clasp2 zebrafish construct for the rescue experiments. S.B. performed western blots and all experiments with ESCs. C.R. performed c-Kit mean intensity measurements. I.S. designed c-Kit intensity measurement and helped with statistical analyses. J.M. performed runx1 WISH. C.R. analyzed and interpreted the experiments with help from all authors. C.R. created the figures and wrote the paper mostly with help from A.K. and N.G. All authors commented on the manuscript. The authors declare no competing financial interests. Funding Information: The authors thank the present and past lab members for helpful discussions and technical help, especially Jean-Charles Boisset, Carla Kroon, Trung Bui, and Rutger Wielink. We also thank the Jeroen den Hertog and Jeroen Bakkers labs for providing reagents, zebrafish lines, and expertise in genome editing and the Catherine Rabouille lab for antibodies. We thank Romualdo Ciau-Uitz (Prof. Roger Patient’s Laboratory) for kindly providing ISH probes. We thank Umut Akinci and Alex Maas (Department of Cell Biology, Erasmus MC) for derivation of the Clasp2 −/− ESCs. We thank the Animal Facility for mouse and zebrafish care and the Optical Imaging Center for confocal microscope access (Hubrecht Institute). We thank Reinier van der Linden (Hubrecht Institute) for help with cell sorting. The authors acknowledge the use of Servier Medical Art image bank to partly create the graphical abstract. This work was supported in the C.R. and N.G. labs by a Landsteiner Stichting voor Bloedtransfusieresearch ( LSBR 1025 ), in the N.G. lab by an NWO-TTW grant ( 15511 ), and in the C.R. lab by a European Research Council grant (ERC project number 220-H75001EU/HSCOrigin-309361 ), a TOP subsidy from NWO/ZonMw ( 912.15.017 ), and an UMC Utrecht “Regenerative Medicine & Stem Cells” priority research program. Publisher Copyright: © 2022 The Author(s)
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