Effects of Diabetes on Retinal Protein Lysine Malonylation

Purpose: The addition of short chain acyl groups to lysines is emerging as key post-translational modifications (PTM) linking metabolite levels to metabolic enzyme activities. Malonyl-lysine (Kmal) targets and inhibits glycolysis and beta-oxidation enzymes. Recently, formation of Kmal was suggested to act as a sink for mitochondrial malate, preventing its inhibition of succinate dehydrogenase in the electron transport chain (ETC). Using the db/db T2DM mouse model, we recently found that diabetes increases retinal glycolysis and beta-oxidation metabolite levels, whereas malate was the only TCA cycle metabolite significantly increased. Metabolic flux measurement with ¹³C-labeled glucose also demonstrated increased enrichment of labeled malate. We therefore compared the relative levels of Kmal-modified proteins in retinas from diabetic mice as well as in postmortem retinas from diabetic patients to determine if this PTM is altered by diabetes.

Methods: Immunofluorescence (IF) with antibodies specific to acetyl-lysine, to succinyl-lysine and to Kmal was used to probe retinal sections from BKS-db/db mice and BKS-db/+ controls (n=9/group) at 24 weeks of age. Kmal modifications were also examined in mouse retinas from mice made diabetic with streptozotocin (STZ) and controls (n=6/group) and a limited number of retinal sections from postmortem eyes of diabetic (n=4) and control (n=2) patients. Antibodies to markers of retinal ganglion and glial cells were used to co-localize Kmal-modified proteins.

Results: Only Kmal IF intensity was significantly reduced by approximately 30% in both db/db and STZ diabetic mouse retinas. Kmal IF exhibited a 25% reduction in human diabetic retinas. Kmal-modified proteins were concentrated in the ganglion cell layer and largely coincided with Müller glia endfeet and/or astrocytes.

Conclusions: The results suggest that protein malonylation in glial cells is diminished in the diabetic retina, which may increase glycolysis and beta-oxidation while inhibiting the ETC.