Atherosclerosis-associated cardiovascular disease is one of the main causes of death and disability among patients with diabetes mellitus. However, little is known about the impact of S-nitrosylation in diabetes-accelerated atherosclerosis. Here, we show increased levels of S-nitrosylation of guanine nucleotide-binding protein G(i) subunit alpha-2 (SNO-GNAI2) at Cysteine 66 in coronary artery samples from diabetic patients with atherosclerosis, consistently with results from mice. Mechanistically, SNO-GNAI2 acted by coupling with CXCR5 to dephosphorylate the Hippo pathway kinase LATS1, thereby leading to nuclear translocation of YAP and promoting an inflammatory response in endothelial cells. Furthermore, Cys-mutant GNAI2 refractory to S-nitrosylation abrogated GNAI2-CXCR5 coupling, alleviated atherosclerosis in diabetic mice, restored Hippo activity, and reduced endothelial inflammation. In addition, we showed that melatonin treatment restored endothelial function and protected against diabetes-accelerated atherosclerosis by preventing GNAI2 S-nitrosylation. In conclusion, SNO-GNAI2 drives diabetes-accelerated atherosclerosis by coupling with CXCR5 and activating YAP-dependent endothelial inflammation, and reducing SNO-GNAI2 is an efficient strategy for alleviating diabetes-accelerated atherosclerosis.

BACKGROUND: Aortic aneurysm and aortic dissection (AAD) are life-threatening vascular diseases, with endothelium being the primary target for AAD treatment. Protein S-sulfhydration is a newly discovered posttranslational modification whose role in AAD has not yet been defined. This study aims to investigate whether protein S-sulfhydration in the endothelium regulates AAD and its underlying mechanism. METHODS: Protein S-sulfhydration in endothelial cells (ECs) during AAD was detected and hub genes regulating homeostasis of the endothelium were identified. Clinical data of patients with AAD and healthy controls were collected, and the level of the cystathionine γ lyase (CSE)/hydrogen sulfide (H 2 S) system in plasma and aortic tissue were determined. Mice with EC-specific CSE deletion or overexpression were generated, and the progression of AAD was determined. Unbiased proteomics and coimmunoprecipitation combined with mass spectrometry analysis were conducted to determine the upstream regulators of the CSE/H 2 S system and the findings were confirmed in transgenic mice. RESULTS: Higher plasma H 2 S levels were associated with a lower risk of AAD, after adjustment for common risk factors. CSE was reduced in the endothelium of AAD mouse and aorta of patients with AAD. Protein S-sulfhydration was reduced in the endothelium during AAD and protein disulfide isomerase (PDI) was the main target. S-sulfhydration of PDI at Cys343 and Cys400 enhanced PDI activity and mitigated endoplasmic reticulum stress. EC-specific CSE deletion was exacerbated, and EC-specific overexpression of CSE alleviated the progression of AAD through regulating the S-sulfhydration of PDI. ZEB2 (zinc finger E-box binding homeobox 2) recruited the HDAC1-NuRD complex (histone deacetylase 1–nucleosome remodeling and deacetylase) to repress the transcription of CTH , the gene encoding CSE, and inhibited PDI S-sulfhydration. EC-specific HDAC1 deletion increased PDI S-sulfhydration and alleviated AAD. Increasing PDI S-sulfhydration with the H 2 S donor GYY4137 or pharmacologically inhibiting HDAC1 activity with entinostat alleviated the progression of AAD. CONCLUSIONS: Decreased plasma H 2 S levels are associated with an increased risk of aortic dissection. The endothelial ZEB2-HDAC1-NuRD complex transcriptionally represses CTH , impairs PDI S-sulfhydration, and drives AAD. The regulation of this pathway effectively prevents AAD progression.

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Protein S-nitrosylation plays an important role in the progression of cardiovascular diseases. eNOS can be S-nitrosylated in endothelial cells, and this modification reversibly attenuates enzyme activity. Under physiological conditions, eNOS directly interacts with β‑catenin. However, whether and how eNOS S-nitrosylation regulates the β‑catenin signal pathway and participates in endothelial dysfunction remains unknown. Here, we show that OxLDL induces the S-nitrosylation of eNOS, which enhances the interaction between eNOS and β‑catenin, transcriptional activity of β‑catenin, cell migration and adhesion molecule expression in endothelial cells. In addition, these effects are partially abolished after eNOS is mutated at Cys94 and Cys99, but not Cys441, in endothelial cells. Furthermore, OxLDL increases iNOS expression. The specific iNOS inhibitor 1400 W decreases eNOS S-nitrosylation and the association of eNOS and β‑catenin, thereby blocking the β‑catenin signal pathway to alleviate OxLDL-induced endothelial dysfunction. Taken together, OxLDL induces eNOS S-nitrosylation at Cys94 and Cys99 via an iNOS-dependent manner, which may increase β‑catenin activation and trigger endothelial injury. This study describes a novel mechanism of endothelial dysfunction.