Phospholamban Acetylation Enhances Cardiomyocyte Calcium Cycling Under Conditions of High-Fat Feeding AHA Conference Repository

American Heart Association

Final ID: Mo069

Phospholamban Acetylation Enhances Cardiomyocyte Calcium Cycling Under Conditions of High-Fat Feeding

Abstract Body:
BACKGROUND:Diet-induced obesity (DIO) is an increasing driver of cardiovascular disease and cardiac dysfunction. The breakdown of excessive fat and sugar intake increases the presence of intracellular acetyl-CoA, which promotes the acetylation of available lysines, altering the stability and activity of these proteins. While numerous proteins involved in cardiac health have been identified as targets of acetylation, the impact of acetylation on calcium handling sarcoplasmic reticulum (SR) proteins, particularly the SERCA2a regulator phospholamban, remains poorly understood.
HYPOTHESIS: Phospholamban acetylation is upregulated by exposure to high fat conditions, in turn driving SERCA2a activity and promoting faster calcium SR reuptake.
APPROACH: Left ventricle (LV) tissue from DIO mice and controls, and human LV tissue from control and diabetic patients, was collected and analyzed using immunoblotting and acetylation was quantified by immunoprecipitation with acetyl-lysine antibodies.Isolated working hearts from DIO and control mice were assessed before and after treatment with acute HDAC activator ITSA-1 and SIRT1 activator SRT1720 to promote deacetylation. Adult ventricular cardiomyocytes were isolated from DIO and control mice, and DIO cells were incubated in media containing excess lipid substrates (1% Lipid Mix 1, Millipore Sigma) and ITSA-1 orSRT1720. Changes in stimulated calcium dynamics were assessed using Fura-2AM and Fluo-4AM imaging. Acetylmimetic/acetylation incompetent PLN mutants were transfected into cultured cells to evaluate SERCA2a binding, oligomerization, and FRET efficiency.
RESULTS: Phospholamban acetylation significantly increased in LV tissue from both DIO mice and human diabetic patients. Acute treatment of isolated hearts and isolated cardiomyocytes from DIO mice with HDAC and SIRT1 activators significantly reduced contractility, relaxation, and SR calcium reuptake rates. Mutations in PLN mimicking or preventing lysine acetylation altered oligomerization and SERCA2a binding.
CONCLUSIONS: Obesity increases the acetylation of phospholamban and enhances the reuptake of calcium into the SR, while deacetylation reverses these effects. These data directly tie nutritive status to a novel phospholamban post-translational modification that may directly impact myocardial calcium handling and contractile function.

Mushala, Bellina ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )
Cho, Ellen ( Loyola University , Chicago , Illinois , United States )
Stoner, Michael ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )
Gibson, Greg ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )
Sembrat, John ( UPMC , Pittsburgh , Pennsylvania , United States )
Bugga, Paramesha ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )
Scott, Iain ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )
Robia, Seth ( Loyola University , Chicago , Illinois , United States )
Manning, Janet ( University of Pittsburgh , Pittsburgh , Pennsylvania , United States )

Author Disclosures:

Bellina Mushala: