Genetic deletion of Cdc42GAP reveals a role of Cdc42 in erythropoiesis and hematopoietic stem/progenitor cell survival, adhesion, and engraftment.
ABSTRACT Rho family GTPases are key signal transducers in cell regulation. Although a body of literature has implicated the Rho family members Rac1 and Rac2 in multiple hematopoietic-cell functions, the role of Cdc42 in hematopoiesis remains unclear. Here we have examined the hematopoietic properties and the hematopoietic stem/progenitor cell (HSP) functions of gene-targeted mice carrying null alleles of cdc42gap, a negative regulator of Cdc42. The Cdc42GAP-/- fetal liver and bone marrow cells showed a 3-fold increase in Cdc42 activity but normal Rac and RhoA activities, indicating that Cdc42GAP knockout resulted in a gain of Cdc42 activity in the hematopoietic tissues. Cdc42GAP-/- mice were anemic. The cellularity of fetal liver and bone marrow, the number and composition percentage of HSPs, and the erythroid blast-forming unit and colony-forming unit (BFU-E/CFU-E) activities were significantly reduced in the homozygous mice. The decrease in HSP number was associated with increased apoptosis of the Cdc42GAP-/- HSPs and the activation of JNK-mediated apoptotic machinery. Moreover, homozygous HSPs showed impaired cortical F-actin assembly, deficiency in adhesion and migration, and defective engraftment. These results provide evidence that Cdc42 activity is important for erythropoiesis and for multiple HSP functions, including survival, adhesion, and engraftment.
SourceAvailable from: Affif ZaccariaInternational Journal of Nanomedicine 03/2015; 2015(10):1869. DOI:10.2147/IJN.S70503 · 4.20 Impact Factor
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ABSTRACT: ARAP3 is a GTPase-activating protein (GAP) that inactivates Arf6 and RhoA small GTPases. ARAP3 deficiency in mice causes a sprouting angiogenic defect resulting in embryonic lethality by E11. Mice with an ARAP3 R302,303A mutation (Arap3KI/KI) that prevents activation by phosphoinositide-3-kinase (PI3K) have a similar angiogenic phenotype, although some animals survive to adulthood. Here, we report that hematopoietic stem cells (HSCs) from rare adult Arap3KI/KI bone marrow are compromised in their ability to reconstitute recipient mice and to self-renew. To elucidate the potential cell-autonomous and non-cell-autonomous roles of ARAP3 in hematopoiesis, we conditionally deleted Arap3 in hematopoietic cells and in several cell types within the HSC niche. Excision of Arap3 in hematopoietic cells using Vav1-Cre does not alter the ability of ARAP3-deficient progenitor cells to proliferate and differentiate in vitro or ARAP3-deficient HSCs to provide multi-lineage reconstitution and to undergo self-renewal in vivo. Thus, our data suggest that ARAP3 does not play a cell-autonomous role in HSPCs. Deletion of Arap3 in osteoblasts and mesenchymal stromal cells using Prx1-Cre resulted in no discernable phenotypes in hematopoietic development or HSC homeostasis in adult mice. In contrast, deletion of Arap3 using vascular endothelial cadherin (VEC or Cdh5)-driven Cre resulted in embryonic lethality, however HSCs from surviving adult mice were largely normal. Reverse transplantations into VEC-driven Arap3 conditional knockout mice revealed no discernable difference in HSC frequencies or function in comparison to control mice. Taken together, our investigation suggests that despite a critical role for ARAP3 in embryonic vascular development, its loss in endothelial cells minimally impacts HSCs in adult bone marrow.PLoS ONE 12/2014; 9(12):e116107. DOI:10.1371/journal.pone.0116107 · 3.53 Impact Factor