Rac1 functions downstream of miR-142 in regulation of erythropoiesis.

Hematopoietic-specific miR-142 is a critical regulator of various blood cell lineages including CD4 dendritic cells and platelet biogenesis in megakaryocytes. Furthermore, we recently reported that miR-142 is required in order to maintain the biconcave shape of erythrocytes, their structural resilience and lifespan, through a mechanism that involves actin filament homeostasis. Here, we used a mouse loss of function allele to characterize a new axis, where miR-142 functions upstream of Rac1 in regulating erythropoiesis. The expression of the two mature miRNAs, miR-1423p and miR-142-5p, emerging from the miR-142 precursor (pre-miR-142) is completely nullified in the hematopoietic lineages of the mouse model that we developed, including in circulating blood cells (Online Supplementary Figure S1A) and in red blood cells (Online Supplementary Figure S1B). miR-142-deficient mice suffer from mild anemia that is associated with a shorter lifespan of adult red blood cells and with reticulocytosis. Therefore, we hypothesized that compensatory erythropoiesis takes place at the bone marrow (BM). Unexpectedly, macroscopic examination of femurs and of freshly isolated BMs revealed erythroid hypoplasia in the BM in miR-142 animals, relative to controls (Figure 1A). The reduced BM cellularity is probably not due to fibrosis, since reticulin staining was distributed in the expected pattern, in both wild-type (WT) and miR-142 bones (Online Supplementary Figure S2). We then characterized erythropoiesis directly by erythroid precursor cytometry, using antibodies against the surface markers CD71 and TER119, which showed impairment in the normoblast series (Figure 1B and C). Therefore, erythropoiesis is impaired in miR-142 knockout animals. miR-142 mice exhibit splenomegaly and disorganized spleen parenchyma; smaller white pulp nodules and red pulp expansion (Online Supplementary Figure S3A and B) are consistent with our previous report. In theory,