SIRT1‐mediated Lysine Crotonylation is Involved in the Regulation of CaMKII Activity in Myocardium

Abstract

ObjectivesPost‐translational modifications (PTMs) can modify proteins and regulate their functions. Lysine crotonylation is a novel protein PTM that can be regulated by silent mating type information regulation 2 homolog 1 (SIRT1). Ca2+/calmodulin‐dependent protein kinase II (CaMKII) plays a significant role in mediating cellular responses to external signaling molecules. Little is known regarding the effect of SIRT1 on CaMKII crotonylation and the impact of crotonylation on the activity of this enzyme.MethodsCre‐Loxp technology was used to generate conditional cardiac‐specific SIRT1 knockout mice (SIRT1 KO). Protein crotonylation modification was studied by PTMomics. CaMKII activity was determined by ELISA. Western blot and co‐immunoprecipitation (CO‐IP) were performed to examine the expression and crotonylation of CaMKIIResultsPTMomics showed changes in protein crotonylation profiles in the myocardium of cardiac‐specific SIRT1 knockout mice. Crotonylation of CaMKII was significantly increased at lysine 301 and the enhanced CaMKII crotonylation was further confirmed by CO‐IP (2.37±0.25 in SIRT1 KO vs. 0.96±0.10 in wild‐type and 1.04±0.10 in SIRT1fl/fl mice, p<0.01). Cardiac‐specific knockout of SIRT1 did not affect the protein expression of CaMKII whereas significantly increased CaMKII activity (2.75±0.17 m units/μL in SIRT1 KO vs. 2.10±0.13 m units/μL in wild‐type and 1.98±0.17 m units/μL in SIRT1fl/fl mice, p<0.05). CO‐IP showed no physical interaction between SIRT1 and CaMKII.ConclusionsSIRT1 deficiency increases CaMKII activity in myocardium at least partially through enhancing lysine crotonylation of CaMKII, which advances our understanding of the role played by PTM in the regulation of CaMKII.