Regulation of Myocardial Ketone Oxidative Proteins by Increased O-GlcNAcylation

An adult heart exhibits tremendous metabolic flexibility by utilizing a variety of substrates to meet its constant fuel demand. However, this metabolic flexibility is lost during overt diabetes. Moreover, heart mitochondria exhibit a low rate of ketone oxidation in type 1 diabetes. However, the mechanism of this reduction and regulation of the ketolytic pathway in a diabetic heart is incompletely understood. Recently, O ‐linked attachment of N‐acetyl‐glucosamine ( O ‐GlcNAc) to proteins has been recognized as a key glucose‐induced posttranslational modification (PTM) in mediating the adverse effects of diabetes in the cardiovascular system. We therefore sought to test the hypothesis that modulation of transcriptional and/or posttranslational mechanisms via increased protein O ‐GlcNAcylation may induce regulatory changes in the myocardial ketolytic machinery in a diabetic heart. Our findings from the hearts of streptozotocin‐induced diabetic mice confirmed that diabetes promotes significant (P < 0.05) transcriptional suppression of two important ketolytic genes: succinyl‐CoA:3‐oxoacid CoA transferase (SCOT; encoded by Oxct1 ) and 3‐hydroxybutyrate dehydrogenase, type 1 (BDH1; encoded by Bdh1 ). In contrast we found an induction of a ketone synthesis gene, 3‐hydroxy‐3‐methylglutaryl‐Coenzyme A synthase 2 (HMGCS2; encoded by Hmgcs2 ). To dissect out the glucose contribution to the changes seen in the diabetic heart, a transgenic mouse with cardiac‐restricted expression of the glucose transporter 4 (GLUT4; mG4H) was used. Interestingly, mG4H alone was sufficient to reduce cardiac Oxct1 expression of SCOT mRNA and protein levels. This transgene‐specific suppression was maintained even in the diabetic myocardium suggesting that glucose may modulate myocardial ketone body catabolic machinery by multiple mechanisms independent of other changes associated with diabetes. To establish the role of O ‐GlcNAcylation, AC16 human cardiomyocytes were treated with Thiamet G (TMG), an O ‐GlcNAcase (OGA) enzyme inhibitor that causes increased protein O ‐GlcNAcylation. TMG‐treatment increased total protein O‐ GlcNAcylation by 2.3‐fold and caused significant reduction in SCOT protein levels (P < 0.01). To further support these findings, a second mouse model was used that overexpressed an OGA‐inactive splice variant of the O ‐GlcNAcase gene (dnOGAh), specifically in cardiac muscle using the α‐myosin heavy chain promoter. Interestingly, cardiac SCOT expression was reduced by 39% in dnOGAh mice following increased cardiac protein O ‐GlcNAcylation supporting a direct role for O ‐GlcNAcylation in the regulation of ketolytic machinery. Together, these results indicate that enhanced glucose delivery into the myocardium may directly regulate myocardial ketone utilization via increased protein O ‐GlcNAcylation. Ongoing in vitro and in vivo studies focus on functional relevance of observed changes that will identify a novel role for cardiac protein O ‐GlcNAcylation, thereby establishing a unique cross‐talk between glucose and ketone metabolism in the heart. Support or Funding Information NIH R00 HL111322, R00 HL111322‐04S1, and U24 DK076169 and Diabetic Complications Consortium (DiaComp) to A.R.W.