Post-translational modifications of autophagy-related (ATG) genes are necessary to modulate their functions. However, ATG protein methylation and its physiological role have not yet been elucidated. The methylation of non-histone proteins by SETD7, a SET domain-containing lysine methyltransferase, is a novel regulatory mechanism to control cell protein function in response to various cellular stresses. Here we present evidence that the precise activity of ATG16L1 protein in hypoxia/reoxygenation (H/R)-treated cardiomyocytes is regulated by a balanced methylation and phosphorylation switch. We first show that H/R promotes autophagy and decreases SETD7 expression, whereas autophagy inhibition by 3-MA increases SETD7 level in cardiomyocytes, implying a tight correlation between autophagy and SETD7. Then we demonstrate that SETD7 methylates ATG16L1 at lysine 151 while KDM1A/LSD1 (lysine demethylase 1A) removes this methyl mark. Furthermore, we validate that this methylation at lysine 151 impairs the binding of ATG16L1 to the ATG12-ATG5 conjugate, leading to inhibition of autophagy and increased apoptosis in H/R-treated cardiomyocytes. However, the cardiomyocytes with shRNA-knocked down SETD7 or inhibition of SETD7 activity by a small molecule chemical, display increased autophagy and decreased apoptosis following H/R treatment. Additionally, methylation at lysine 151 inhibits phosphorylation of ATG16L1 at S139 by CSNK2 which was previously shown to be critical for autophagy maintenance, and vice versa. Together, our findings define a novel modification of ATG16L1 and highlight the importance of an ATG16L1 phosphorylation-methylation switch in determining the fate of H/R-treated cardiomyocytes.
[1]Affiliated Shanghai Univ Med & Hlth Sci, Jiading Dist Cent Hosp, Dept Cardiol, Shanghai, Peoples R China;
[2]Zunyi Med Univ, Longju Med Res Ctr, Key Lab Basic Pharmacol, Minist Educ, Zunyi, Peoples R China;
[3]Rutgers State Univ, Rutgers Canc Inst New Jersey, New Brunswick, NJ USA;
[4]Huazhong Univ Sci & Technol, Union Hosp, Tongji Med Coll,Minist Educ, Lab Cardiovasc Immunol,Key Lab Biol Targeted Ther, Wuhan, Hubei, Peoples R China;
[5]Fudan Univ, Minhang Hosp, Dept Cardiol, Shanghai, Peoples R China;
[6]Huazhong Univ Sci & Technol, Tongji Hosp, Tongji Med Coll, Dept Cardiol, Wuhan, Peoples R China;
[7]Harbin Med Univ, Affiliated Hosp 1, Dept Cardiol, Harbin, Peoples R China;
[8]Zunyi Med Univ, Key Lab Basic Pharmacol, Minist Educ, Zunyi, Peoples R China;
[9]Affiliated Hosp Zunyi Med Univ, Dept Cardiovasc Surg, Zunyi, Peoples R China;
[10]Affiliated Hosp Zunyi Med Univ, Dept Anesthesia, Zunyi, Peoples R China;
[11]State Univ New Jersey, Rutgers Univ, Univ Hosp, Robert Wood Johnson Med Sch,Dept Surg, New Brunswick, NJ USA
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