OGT-Mediated KEAP1 Glycosylation Accelerates NRF2 Degradation Leading to High Phosphate-Induced Vascular Calcification in Chronic Kidney Disease.

Tian-Hua Xu,Yue Li,Li Yao,Yinke Du,Zitong Sheng,Xiaobo Qiu,Binyao Tian

doi:10.3389/fphys.2020.01092

Abstract

Unraveling the complex regulatory pathways that mediate the effects of phosphate on vascular smooth muscle cells (VSMCs) may provide novel targets and therapies to limit the destructive effects of vascular calcification (VC) in patients with chronic kidney disease (CKD). Our previous studies have highlighted several signaling networks associated with VSMC autophagy, but the underlying mechanisms remain poorly understood. Thereafter, the current study was performed to characterize the functional relevance of O-linked N-acetylglucosamine (GlcNAc) transferase (OGT) in high phosphate-induced VC in CKD settings. We generated VC models in 5/6 nephrectomized rats in vivo and VSMC calcification models in vitro. Artificial modulation of OGT (knockdown and overexpression) was performed to explore the role of OGT in VSMC autophagy and VC in thoracic aorta, and in vivo experiments were used to substantiate in vitro findings. Mechanistically, co-immunoprecipitation (Co-IP) assay was performed to examine interaction between OGT and kelch like ECH associated protein 1 (KEAP1), and in vivo ubiquitination assay was performed to examine ubiquitination extent of nuclear factor erythroid 2-related factor 2 (NRF2). OGT was highly expressed in high phosphate-induced 5/6 nephrectomized rats and VSMCs. OGT silencing was shown to suppress high phosphate-induced calcification of VSMCs. OGT enhances KEAP1 glycosylation and thereby results in degradation and ubiquitination of NRF2, concurrently inhibiting VSMC autophagy to promote VSMC calcification in 5/6 nephrectomized rats. OGT inhibits VSMC autophagy through the KEAP1/NRF2 axis and thus accelerates high phosphate-induced VC in CKD.

Highlights

Chronic kidney disease (CKD) is typically accompanied by destructive complications such as accumulating atherosclerotic plaque calcification and medial calcifications (Neven and D’Haese, 2011)
We attempted to investigate the function of O-linked N-acetylglucosamine (GlcNAc) transferase (OGT) in high phosphate-induced vascular calcification (VC) in chronic kidney disease (CKD)
Elevated expression of OGT was identified in the above-mentioned rat models through immunohistochemical detection (Figure 1E), relative to sham-operated rats that showed no differences from the normal rats

Summary

Introduction

Chronic kidney disease (CKD) is typically accompanied by destructive complications such as accumulating atherosclerotic plaque calcification and medial calcifications (Neven and D’Haese, 2011). Cardiovascular complications are recognized as the leading cause of death in patients with CKD (Mizobuchi et al, 2009). High serum phosphate concentrations are known to commonly lead to increased risk of cardiovascular diseases (CVDs) and OGT/KEAP1/NRF2 Axis in VC of CKD exacerbation of CKD (Yoon et al, 2017). Recent evidence has demonstrated that high phosphate exposure or phosphate loading directly results in endothelial and vascular dysfunction in blood vessels, a contributor to cardiovascular risk in CKD (Stevens et al, 2015). Our team has previously provided evidence verifying the ameliorative role of autophagy induction of VSMCs in hyperphosphatemia-induced VC, which implicates the nuclear factor erythroid 2-related factor 2 (NRF2)/ARE pathway (Yao et al, 2017). Further efforts are still needed to explore the molecular mechanism associated with VSMC autophagy in VC in CKD setting

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