Integrative proteomics reveals principles of dynamic phosphosignaling networks in human erythropoiesis

Human erythropoiesis is an exquisitely controlled multistep developmental process, and its dysregulation leads to numerous human diseases. Transcriptome and epigenome studies provided insights into system‐wide regulation, but we currently lack a global mechanistic view on the dynamics of proteome and post‐translational regulation coordinating erythroid maturation. We established a mass spectrometry (MS)‐based proteomics workflow to quantify and dynamically track 7,400 proteins and 27,000 phosphorylation sites of five distinct maturation stages of in vitro reconstituted erythropoiesis of CD34(+) HSPCs. Our data reveal developmental regulation through drastic proteome remodeling across stages of erythroid maturation encompassing most protein classes. This includes various orchestrated changes in solute carriers indicating adjustments to altered metabolic requirements. To define the distinct proteome of each maturation stage, we developed a computational deconvolution approach which revealed stage‐specific marker proteins. The dynamic phosphoproteomes combined with a kinome‐targeted CRISPR/Cas9 screen uncovered coordinated networks of erythropoietic kinases and pinpointed downregulation of c‐Kit/MAPK signaling axis as key driver of maturation. Our system‐wide view establishes the functional dynamic of complex phosphosignaling networks and regulation through proteome remodeling in erythropoiesis.