Effects of diabetes on retinal protein lysine malonylation

Abstract

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. Lysine malonylation targets and inhibits glycolysis and beta-oxidation enzymes. Recently, formation of malonyl-lysine (MalK) 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 caused increases in retinal glycolysis and beta-oxidation metabolite levels, whereas malate was the only TCA metabolite significantly increased. Metabolic flux measurement with C¹³-labeled glucose also demonstrated increased enrichment of labeled malate. We therefore compared the relative levels of MalK-modified proteins in retinas from type 1 and type 2 diabetic mice to determine if this PTM is altered by diabetes.

Methods : Immunofluorescence (IF) with antibodies specific to acetyl-lysine (AcK), succinyl-lysine (SucK) and MalK was used to examine retinal sections from BKS-db/db mice and BKS-db/+ controls (n=9/group) at 24 weeks of age. These modifications were also examined in retinas from mice made diabetic with streptozotocin (STZ) and controls (n=10/group) after 16 weeks of diabetes. Antibodies to protein markers for retinal ganglion cells (brain-specific homeobox/POU domain protein 3A), Müller cells (glutamine synthetase) and astrocytes (glial fibrillary acidic protein) were used to co-localize MalK-modified proteins.

Results : MalK-modified proteins were concentrated in the retinal ganglion cell layer (GCL) with the highest IF intensity coinciding with Müller glia endfeet and astrocytes. MalK IF intensity in all retinal layers was significantly reduced by approximately 30% in both db/db and STZ diabetic mouse retinas. MalK IF within the GCL of diabetic mice was similarly reduced. AcK and SucK IF intensities were not altered by diabetes.

Conclusions : The results suggest that protein malonylation is high in normal retina glial cells and is diminished in the diabetic retina. Loss of MalK PTM on metabolic enzymes could increase glycolysis and beta-oxidation while inhibiting the ETC in the diabetic retina. Reversal of protein malonylation may be an adaptation to the changes in metabolite delivery caused by diabetes and/or may lead to retinal dysfunction that contributes to diabetic retinopathy.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.