Cytoskeletal rearrangements and SRC signalling defects controlled by PP1β drive impaired megakaryocyte maturation and platelet production

Abstract Platelet production is a highly regulated process involving coordinated signaling and cytoskeletal remodeling. While thrombopoietin-driven pathways have been well characterized, the contribution of serine/threonine phosphatases remains poorly explored. In this study, we interrogated the specific function of the phosphatase PP1β in megakaryocyte development using a lineage-specific conditional knockout mouse model by crossing Ppp1cb-floxed mice with Pf4-Cre transgenic mice. Mice lacking PP1β in the megakaryocyte lineage exhibited severe macrothrombocytopenia, accompanied by hallmark features of bone marrow pathology, including myelofibrosis, osteosclerosis, and splenic enlargement. The depletion of PP1β let to an accumulation of smaller megakaryocytes in the bone marrow, which displayed abnormal ultrastructure and disorganized cytoplasm with impaired demarcation membrane system formation. In vitro assays confirmed elevated ploidy, changes in the expression of surface glycoproteins critical for platelet function defective proplatelet formation, and lower platelet lifespan. Notably, these main findings were also recapitulated in vivo using intravital imaging assays, further validating the physiological relevance of the observed defects. Mechanistically, we observed increased phosphorylation of myosin light chain 2 (Ser19), consistent with enhanced actomyosin contractility and stress fiber assembly. Surprisingly, canonical thrombopoietin signaling via Akt and STAT family members remained unaltered, while phosphorylation of Src family kinases at their activation loop was markedly diminished. These data suggest that PP1β modulates both cytoskeletal tension and tyrosine kinase signaling to support megakaryocyte maturation and platelet release. Together, our findings position PP1β as an essential regulator of megakaryopoiesis, acting through control of myosin II activity and Src kinase signaling, highlighting a distinct, non-redundant role for this phosphatase in hematopoiesis.