Megakaryocyte morphogenesis employs a "hypertrophy-like" developmental program that is dependent on P-TEFb kinase activation and cytoskeletal remodeling. P-TEFb activation classically occurs by a feedback-regulated process of signal-induced, reversible release of active Cdk9-cyclin T modules from large, inactive 7SK small nuclear ribonucleoprotein particle (snRNP) complexes. Here, we have identified an alternative pathway of irreversible P-TEFb activation in megakaryopoiesis that is mediated by dissolution of the 7SK snRNP complex. In this pathway, calpain 2 cleavage of the core 7SK snRNP component MePCE promoted P-TEFb release and consequent upregulation of a cohort of cytoskeleton remodeling factors, including α-actinin-1. In a subset of human megakaryocytic leukemias, the transcription factor GATA1 undergoes truncating mutation (GATA1s). Here, we linked the GATA1s mutation to defects in megakaryocytic upregulation of calpain 2 and of P-TEFb-dependent cytoskeletal remodeling factors. Restoring calpain 2 expression in GATA1s mutant megakaryocytes rescued normal development, implicating this morphogenetic pathway as a target in human leukemogenesis.