Non-muscle Myosin II drives vesicle loss during human reticulocyte maturation

The process of reticulocyte maturation into fully mature erythrocytes that occurs in circulation is known to be characterised by a complex interplay between loss of cell surface area and volume, removal of remnant cell organelles and redundant proteins, and highly selective membrane and cytoskeletal remodelling. However, the mechanisms that underlie and drive these maturational processes in vivo are currently poorly understood and, at present, reticulocytes derived through in vitro culture fail to undergo the final transition to erythrocytes. Here, we use high-throughput proteomic methods to highlight differences between erythrocytes, cultured and endogenous reticulocytes. We identify a cytoskeletal protein, non-muscle myosin IIA (NMIIA) as exhibiting differential abundance and phosphorylation status between reticulocytes and erythrocytes and localize it in the proximity of autophagosomal vesicles. An ex vivo circulation system was developed to simulate the mechanical shear component of circulation and demonstrated that mechanical stimulus is necessary, but insufficient for reticulocyte maturation. Using this system in concurrence with NMII inhibition, we demonstrate involvement of NMIIA in reticulocyte remodelling and propose a previously undescribed mechanism of shear stress-responsive vesicle clearance that is crucial for reticulocyte maturation.