Diaphanous-Related Formin Protein mDia2 Regulates Hematopoietic Stem and Progenitor Cell Functions through Srf-Integrins Pathway

Abstract

Distinct mature cell types in mammalian blood system are derived from the Hematopoietic Stem/Progenitor cells (HSPCs), which are multipotent progenitor cells with self-renewing and engraftment ability. Various intrinsic and extrinsic molecules modulate HSPCs' fates to either undergo self-renewal or differentiation. The Diaphanous-related formin protein mDia2, is an effector of Rho GTPases. Previously, we reported that hematopoietic-specific mDia2 knock out mice developed anemia due to ineffective erythropoiesis caused by bi-nucleated erythroblasts. However, how mDia2 is involved in HSPC functions are unknown. Here we show that mDia2 expression is enriched in HSPCs. Although mDia2 was largely dispensable for HSPCs' function at steady-state, hematopoietic-specific inactivation of mDia2 severely impaired the competitive repopulation capacity of HSPCs. After primary bone marrow transplantation, the absolute number of bone marrow HSPCs from mDia2 knockout (KO) transplants was significantly decreased. mDia2 deficient LSK cells also showed skewed differentiation with increased percentage of more differentiated multi-potent progenitor cells (MPP), which was further supported by loss of stem cell quiescence. Under serial transplantation, mice transplanted with mDia2 knockout (KO) bone marrow showed rapid lethality. Importantly, these defects can be rescued by the constitutively active mDia2 mutant mDia2-ΔDAD. Furthermore, mDia2 KO HSPCs showed a marked reduction in the motility ability from bone marrow to the peripheral blood upon G-CSF stimulation. Loss of mDia2 also severely compromised the recovery of the mice from 5-FU-induced myelosuppression.mDia2 functions to nucleate linear actin filaments. We found that loss of mDia2 expression in HSPCs reduced the F-actin levels. This further attenuated the adhesion of HSPCs that was confirmed by adhesion assay and decreased protrusion formation in vitro . We demonstrated that the defect in F-actin formation caused by mDia2 deficiency repressed the transcriptional activity of serum response factor (SRF), which caused down-regulation of SRF target genes such as Acta2, Actb and SRF itself. Ectopic expression of SRF rescued HSPC phenotypes with loss of mDia2. We further identified that the expression levels of several integrins, including Itga2, Itgam (encodes CD11b) , Itagal (encodes CD11a)and Itgb2 (encodes CD18) , were significantly attenuated in mDia2 KO c-Kit+ HSPCs. Chromatin immunoprecipitation (ChIP) sequencing data further revealed intronic SRF binding sites in the genomic loci of Itgam and Itgb2, therefore suggesting Itgam and Itgb2 as new downstream targets of SRF pathway. Our data revealed a novel mDia2-SRF-integrin axis that regulates HSPCs adhesion and engraftment. DisclosuresNo relevant conflicts of interest to declare.