The Role Of PPARγ Acetylation In Transcriptional Regulation Of Klotho And Renal Protection

Klotho is a kidney-enriched anti-aging protein.Decreased Klotho expression closely correlates with the development and progression of chronic kidney diseases.Exogenous Klotho supplementation or upregulation of endogenous Klotho ameliorates chronic kidney injury.Thus,Klotho is a crucial intervention target for chronic kidney injury.PPARy is an important renal protective transcription factor and its functionalities are significantly affected by various post-translational modifications.Modification of PPARy by direct acetylation is a new topic in the study of PPARy function.PPARy acetylation regulates adipocyte transition and lipid metabolism.However,the functional significance of PPARy acetylation in kidney injury is unknown.Klotho is a target gene of PPARy,but whether PPARy acetylation regulates Klotho is not clear.Therefore,in this study we investigate the effect of PPARy acetylation on Klotho expression and chronic kidney injury.Trichostatin A(TSA)induces protein acetylation by histone deacetylase inhibition.We found that TSA significantly alleviated chronic kidney injury and Klotho downregulation in adenine-fed mice.TSA up-regulated Klotho mainly through enhancing PPARy acetylation and promoting PPARy binding to Klotho promoter.We demonstrated that PPARy acetylation regulation of Klotho depends on PPARy acetylations on K240 and K265.TSA reversal of Klotho loss and the renoprotective effect were abrogated in PPARy knockout mice,supporting that PPARy is an essential acetylation target for the Klotho restoration and renal protection.More intriguingly,adenine mouse kidney displayed deregulated HDAC3 up-regulation.Selective HDAC3 inhibitor,RGFP966,effectively alleviated the Klotho loss and chronic renal injuries,whereas the protective effects were largely abolished when Klotho was knocked down by tail vein injection of siRNA,suggesting that aberrant HDAC3 and Klotho loss are the crucial components mediating the renal injuries in adenine mice.Bone abnormalities are a common complication of chronic kidney disease.Kidney-specific Klotho knockout mice exhibit not only renal injuries but also bone disorders.Thus,in this study we explored whether PPARy acetylation regulation of Klotho attenuates chronic kidney injury associated bone complication.We demonstrated adenine mice developed bone abnormalities.The results of ELISA,Western Blot and qPCR showed that bone remodeling factors were significantly abnormal.Bone HE stainings suggested that bone trabecula are thinner and deranged,and the lacuna are increased.The X-ray scanning showed the marked loss of bone mineral density at the distal femurs.MicroCT 3D analysis suggested that distal femur micro-architectures were abnormal,such as less trabecular bone volume fraction(BV/TV),reduced trabecular bone number(Tb.N)and trabecular thickness(Tb.Th),and increased trabecular bone separation(Tb.Sp).Enhancement of Klotho expression by PPARγ acetylation improved the bone abonormalities of adenine mice.Knocking down Klotho expression abolished the protective effects of TSA on bone,suggesting Klotho loss as a key event linking HDAC deregulation to the chronic kidney injury-associated bone disorders.In conclusion,the results from our study represent the first in vivo functional evaluation of PPARy acetylation and the associated Klotho regulation in chronic kidney injury of an adenine mouse model.In addition,our results have therapeutic implications suggesting that HDAC3 inhibition increases PPARy transcriptional activity,retains Klotho and attenuates chronic kidney injury and associated bone abnormalities.