Integrative proteomics reveals principles of dynamic phospho-signaling networks in human erythropoiesis

Human erythropoiesis is exquisitely controlled at multiple levels and its dysregulation leads to numerous human diseases. Despite many functional studies focused on classical regulators, we lack a global, system-wide understanding of post-translational mechanisms coordinating erythroid maturation. Using the latest advances in mass spectrometry (MS)-based proteomics we comprehensively investigate the dynamics of protein and post-translational regulation of in vitro reconstituted CD34+ HSPC-derived erythropoiesis. This quantifies and dynamically tracks 7,400 proteins and 27,000 phosphorylation sites. Our data reveals differential temporal protein expression encompassing most protein classes and numerous post-translational regulatory cascades. Drastic cell surface remodeling across erythropoiesis include numerous orchestrated changes in solute carriers, providing new stage-specific markers. The dynamic phosphoproteomes combined with a kinome-targeting CRISPR/Cas9 screen reveal coordinated networks of erythropoietic kinases and downregulation of MAPK signaling subsequent to c-Kit attenuation as key drivers of maturation. Our global view of erythropoiesis establishes a central role of post-translational regulation in terminal differentiation.