Protein half‐life determines expression of proteostatic networks in podocyte differentiation

Podocytes are highly specialized, epithelial, postmitotic cells, which maintain the renal filtration barrier. When adapting to considerable metabolic and mechanical stress, podocytes need to accurately maintain their proteome. Immortalized podocyte cell lines are a widely used model for studying podocyte biology in health and disease in vitro. In this study, we performed a comprehensive proteomic analysis of the cultured human podocyte proteome in both proliferative and differentiated conditions at a depth of >7000 proteins. Similar to mouse podocytes, human podocyte differentiation involved a shift in proteostasis: undifferentiated podocytes have high expression of proteasomal proteins, whereas differentiated podocytes have high expression of lysosomal proteins. Additional analyses with pulsed stable-isotope labeling by amino acids in cell culture and protein degradation assays determined protein dynamics and half-lives. These studies unraveled a globally increased stability of proteins in differentiated podocytes. Mitochondrial, cytoskeletal, and membrane proteins were stabilized, particularly in differentiated podocytes. Importantly, protein half-lives strongly contributed to protein abundance in each state. These data suggest that regulation of protein turnover of particular cellular functions determines podocyte differentiation, a paradigm involving mitophagy and, potentially, of importance in conditions of increased podocyte stress and damage.-Schroeter, C. B., Koehler, S., Kann, M., Schermer, B., Benzing, T., Brinkkoetter, P. T., Rinschen, M. M. Protein half-life determines expression of proteostatic networks in podocyte differentiation.