Abstract
Retinal neurodegeneration develops early in the course of diabetic retinopathy (DR), and our previous research showed that succinate accumulation results in retinal ganglion cells (RGCs) dysfunction in the retinas of rats with DR. Succinate can enhance lysine succinylation, but the succinylation of DR is not well understood. In this study, we investigated the role of the succinylome in DR and identified the key factor in this process. TMT labeling and LC–MS/MS analysis were combined to quantify the differentially succinylated proteins between vitreous humor (VH) samples from DR and non-DR patients. A total of 74 sites in 35 proteins were differentially succinylated between DR and non-DR vitreous humor samples, among which succinylation of the K108 site of optineurin (OPTN K108su) in the defense response was enriched by GO analysis based on the biological process category. Then, using a streptozotocin (STZ)-induced diabetic rat model, R28 cells and primary rat RGCs (rRGCs), we demonstrated that OPTN underwent lysine succinylation in the retinas of rats with DR and that OPTN K108su mediated autophagic flux blockade under high-glucose (HG) conditions. Sirt5 can desuccinylate OPTN K108su, thus protecting RGCs function from high glucose-induced RGCs autophagic flux blockade in the diabetic retina. Overall, desuccinylation of OPTN is an essential adaptive mechanism for ameliorating autophagic flux blockade in RGCs under DR conditions, and targeting the Sirt5-desuccK108-OPTN axis may thus open an avenue for therapeutic intervention in RCGs dysfunction.
Highlights
Diabetic retinopathy (DR) is a neural and microvascular complication of diabetes mellitus (DM) and remains the leading cause of preventable blindness in working-aged people [1]
Among members of the sirtuin humor from DR patients by MS/MS assay family, Sirt5 has weak deacetylase activity but can catalyze the We combined TMT labeling, highly specific anti-succinyl lysine removal of three acidic lysine modifications, malonylation, pan-antibody enrichment, and LC–MS/MS analysis for the succinylation, and glutarylation [21], and is the only desuccinylase systematic analysis of the succinylome in vitreous humor samples located in the cytoplasm [22, 23]
Lys108 in OPTN is a key site for high glucose-mediated succinylation and autophagic flux blockade in rat RGCs (rRGCs) and R28 cells As endogenous OPTN was succinylated in the vitreous humor and the retina of DR, we explored the modification of OPTN and the role of OPTN K108su using in vitro experiments
Summary
Diabetic retinopathy (DR) is a neural and microvascular complication of diabetes mellitus (DM) and remains the leading cause of preventable blindness in working-aged people [1]. Recent research has demonstrated that neurodegenerative processes develop early in the course of DR, even before the onset of microvascular changes [2–4]. The underlying mechanism of early RGCs dysfunction in DR remains to be clarified, understanding the early effects of diabetes on neural cells in the retina could eventually lead to the early identification and diagnosis of DR. Our previous studies demonstrated that an accumulation of succinate in streptozotocin (STZ)-induced type 1 diabetes mellitus (T1DM) rat retinas promotes RGCs dysfunction in the early stage of DR [6]. Differentially succinylated proteins regulating various biological processes were identified, with particular enrichment of defense response; among these proteins, the succinylation of optineurin (OPTN) interested us and was a previously unreported modification of OPTN
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