O‐GlcNAcylation of CaMKII in Cardiac Myocytes can be Pro‐Arrhythmic

Abstract

We found that high extracellular [glucose] (Hi‐Gluc) increased the activity level of Ca‐Calmodulin dependent protein kinase II (CaMKII) in adult cardiac myocytes, using both traditional biochemical activity measurements as well as a FRET based activation state reporter, Camui. Hi‐Gluc can induce O‐GlcNAcylation (covalent O‐linked β‐D‐N‐acetylglucosamine, GlcNAc) at Ser/Thr residues, and is a ‐translational modification of proteins (often at or near kinase target sites). GlcNAcylation is dynamic and is catalyzed by O‐GlcNAc transferase (OGT) (which is inhibitable upstream by DON) and GlcNAc is removed by O‐GlcNAase (which can be blocked by Thiamet G). Hi‐Gluc‐induced CaMKII activation was blocked by DON, promoted by Thiamet G and isoproterenol (ISO), and also promoted autonomous CaMKII activation (similar to that induced by autophosphorylation and oxidation). Mutation of CaMKII (S279A), adjacent to the known oxidation site prevented Hi‐Gluc –dependent activation, but did not prevent autonomous activation by autophosphorylation or oxidation. Mass spec confirmed S279 as a site of GlcNAcylation, and CaMKII was more highly GlcNAcylated in diabetic hearts and brains (human and rat). Hi‐Gluc caused activation of cardiac myocyte Ca sparks, action potential alterations and cardiac arrhythmias. These effects were prevented by either CaMKII or GlcNAc inhibition (DON) and in genetic CaMKIIδ‐KO mice. This new pathway may be an important pathophysiological synergy between GlcNAcylation and CaMKII signaling relevant in diabetic hyperglycemia.