PPP2R2B Variant Reduces Protein Phosphatase 2A (PP2A) Complexity and Worsens Diabetic Kidney Disease (DKD) Progression

Background: We have previously shown that protein phosphatase 2A (PP2A), a major Ser/Thr phosphatase in the kidney, is involved in regulation of podocyte function in diabetic kidney disease. Podocyte-specific knockout of PP2A catalytic unit leads to more severe podocyte injury and progression of DKD. Here, we report the identification of variants in the PPP2R2B gene, encoding the podocyte-specific regulatory subunit of PP2A, in DKD patients with ESKD. Methods: We carried out clinical studies and genetic analyses, including genome-wide linkage analysis and whole-exome sequencing to identify candidate variants. Transgenic mice models were generated to validate the phenotype of the candidate variant. Functional studies on the disease-associated variant were performed in transgenic mice glomeruli and cultured cells. Results: Several variants of PPP2R2B gene were found to be associated with renal outcome in DKD patients. To help elucidate the underlying mechanism, we generated a knock-in mice model with one of the clinical worst outcome-related PPP2R2B missense variants (p.T222M). We induced diabetes in these knock-in mice and their littermate controls. We found that mice carried Ppp2r2b p.T222M developed worse DKD phenotype such as proteinuria, glomerular hypertrophy, and mesangial expansion, as compared to littermate controls. Crucially, Ppp2r2b p.T222M results in impaired glomerular PP2A activity as measured by PP2A activity assay and phosphorylation of its substrate NF-kB. However, the protein expression level of glomerular Ppp2r2b was not affected by the variant. In addition, using in vitro studies, we showed that PPP2R2B p.T222M variant reduced PP2A complexity, thus altering cellular PP2A activity and downstream phosphorylation of NF-kB. Conclusion: Our findings confirmed that mice carrying Ppp2r2b p.T222M variant developed more severe renal phenotype. We also demonstrated that PPP2R2B p.T222M variant led to impaired glomerular PP2A activity by reducing the complexity of the PP2A holoenzyme, without affecting the protein expression levels. Our study help us better understand the role of PP2A complex in the pathogenesis of human DKD and might provide new insights into therapeutic targets for DKD. Funding: NIDDK Support, Veterans Affairs Support