Epigenetically silencing gluconeogenic enzyme PCK1 by EZH2 promotes renal tubulointerstitial fibrosis

Renal fibrosis is the common pathological pathway of various chronic kidney diseases (CKD) in progression to the end stage of renal failure. The methyltransferase enhancer of zeste homolog 2 (EZH2) has been identified as a therapeutic target to inhibit renal fibrosis. However, the mechanism underlying the role of EZH2 in renal fibrosis is not completely understood. By using EZH2 inhibitor 3-DZNeP and Ezh2 conditional knockout mice, we confirmed the pro-fibrotic effect of EZH2 in unilateral ureteral obstruction (UUO) kidneys. Through RNA sequencing and cleavage under targets and tagmentation (CUT&Tag) sequencing analysis, we found that the phosphoenolpyruvate carboxykinase 1 (PCK1), a critical enzyme in gluconeogenesis, is negatively regulated by EZH2 in fibrotic kidneys, which was further confirmed by quantitative PCR, CUT&Tag and Western blotting analysis in UUO kidneys and renal fibrotic cells. We further showed that deletion or inhibition of EZH2 increased PCK1 expression and attenuated renal fibrosis in two other mouse models. Moreover, the concentration of lactate, a glucogenic metabolic intermediate, was increased in mouse fibrotic kidneys, which was reduced by EZH2 inhibition. We further showed that glucose production was impaired in folic acid induced nephrotic mice and restored by EZH2 inhibition as assessed by pyruvate tolerance test. Importantly, the direct anti-gluconeogenesis effect of EZH2 on renal proximal epithelial cells was shown in human HK2 cells by analysis of gluconeogenic gene expression and production of glucose or lactate. Finally, inhibition of PCK1 by 3-mercaptopropionic acid abrogated the anti-fibrotic effect of EZH2 inhibitor in UUO kidneys. We conclude that EZH2 promotes renal tubulointerstitial fibrosis through epigenetic inhibition of the rate-limiting gluconeogenesis enzyme PCK1. Our study suggests that therapeutic restoration of the impaired renal gluconeogenesis is beneficial to CKD patients. Highlights EZH2 is negatively correlated with the expression of gluconeogenic genes in fibrotic kidneys The expression of gluconeogenic enzyme PCK1 is epigenetically regulated by EZH2 Impaired renal gluconeogenesis in chronic kidney disease can be epigenetically regulated and therapeutically targeted Restoration of renal gluconeogenesis by EZH2 inhibition ameliorates renal tubulointerstitial fibrosis