Membrane budding is a major mechanism of in vivo platelet biogenesis.

How platelets are produced by megakaryocytes in vivo remains controversial despite more than a century of investigation. Megakaryocytes readily produce proplatelet structures in vitro; however, visualization of platelet release from proplatelets in vivo has remained elusive. We show that within the native prenatal and adult environments, the frequency and rate of proplatelet formation is incompatible with the physiological demands of platelet replacement. We resolve this inconsistency by performing in-depth analysis of plasma membrane budding, a cellular process that has previously been dismissed as a source of platelet production. Our studies demonstrate that membrane budding results in the sustained release of platelets directly into the peripheral circulation during both fetal and adult life without induction of cell death or proplatelet formation. In support of this model, we demonstrate that in mice deficient for NF-E2 (the thrombopoietic master regulator), the absence of membrane budding correlates with failure of in vivo platelet production. Accordingly, we propose that membrane budding, rather than proplatelet formation, supplies the majority of the platelet biomass.