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Abstract: TH-PO870

Enhancer of Zeste Homolog-2 (EZH2), a Histone H3 Methyltransferase, Launches Deptor Downregulation for Mesangial Cell (MC) Hypertrophy and Matrix Expansion in Diabetic Nephropathy (DN)

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Das, Falguni, University of Texas Health Science Center, San Antonio, Texas, United States
  • Ghosh-choudhury, Nandini, University of Texas Health Science Center, San Antonio, Texas, United States
  • Kasinath, Balakuntalam S., University of Texas Health Science Center, San Antonio, Texas, United States
  • Ghosh-Choudhury, Goutam, University of Texas Health Science Center, San Antonio, Texas, United States
Background

mTOR has emerged as a centerpiece in pathogenesis of DN. Deptor is an inhibitory component of both mTOR complexes (C1 and C2). We investigated epigenetic control of deptor expression and its relevance to mesangial cell hypertrophy and matrix protein expansion during the progression of DN.

Methods

Human MCs and OVE26 mice were used. Activation-specific antibodies, real time qRT-PCR, immunoblotting, plasmid-derived expression vector and shRNA transfection were employed.

Results

In MCs, 25 mM glucose (HG) decreased expression of deptor mRNA and protein in a time-dependent manner, resulting in increased activation of mTORC1 and C2 as judged by phosphorylation of S6 kinase (T-389) and Akt (S-473), respectively. To determine the mechanism of deptor downregulation, we considered the epigenetic mechanism involving the polycomb repressor complex-2 in which EZH2 promotes the trimethylation (Me3) of histone H3 at K27. HG increased the expression of EZH2 concomitant with increased H3K27 Me3. Deazaneplanocin (DZNep), an inhibitor of EZH2, blocked H3 Me3 and deptor downregulation induced by HG. Also, DZNep inhibited HG-stimulated mTORC1 and C2 activities. Similarly, shRNA against EZH2 inhibited H3K27 Me3, reversed HG-induced deptor inhibition and suppressed mTORC1 and mTORC2 activities. In contrast, expression of EZH2 increased H3 K27 Me3, decreased deptor expression and enhanced mTORC1 and C2 activities, similar to HG treatment. Furthermore, DZNep and shEZH2 significantly inhibited HG-induced MC hypertrophy and expression of fibronectin and plasminogen activator inhibitor-1 (PAI-1). On the other hand, EZH2 increased fibronectin and PAI-1 expression similar to HG. To address the in vivo relevance of our observations, we used OVE26 diabetic mouse. In the renal cortex of these mice, expression of EZH2 was significantly increased concomitant with increased H3K27 Me3, mTORC1 and C2 activities, and fibronectin and PAI-1 expression.

Conclusion

Our results for the first time uncover a precisely tuned balance between deptor suppression by EZH2 and activation of mTORC1 and C2 for HG-induced MC hypertrophy and matrix accumulation. The data lend support for testing EZH2 inhibitors in preclinical model for attenuation of complications of DN.

Funding

  • Veterans Affairs Support