Comprehensive proteomic analysis reveals dynamic phospho-profiling in human early erythropoiesis

Normal early erythropoiesis depends on the precise regulation of protein expression and phosphorylation modification. Dysregulation of protein levels or modification contributes to erythroid disorders. To date, the dynamics of protein phosphorylation profiling across human erythroid development is not fully understood. Here, we characterized quantitative proteomic and phosphoproteomic profiling by tandem mass-tagging technology. We systemically built phospho-expression profiling and expression clusters of 11 414 phosphopeptides for human early erythropoiesis. The standardization methods for multitier integrative analyses revealed multiple functional modules of phosphoproteins (e.g., regulation of the G2/M transition) and active phosphorylated signalling (e.g., cell cycle-related pathways). Our further analysis revealed that CDK family members were the main kinases that phosphorylate substrates in erythroid progenitors and identified that CDK9 played an important role in the proliferation of erythroid progenitors. Collectively, our phosphoproteomic profiling, integrative network analysis and functional studies define landscapes of the phosphoproteome and reveal signalling pathways that are involved in human early erythropoiesis. This study will serve as a valuable resource for further investigations of phosphatase and kinase functions in human erythropoiesis and erythroid-related diseases.