Mitochondrial hyper-acetylation induced by an engineered acetyltransferase promotes cellular senescence.

Protein acetylation plays crucial roles in diverse biological functions, including mitochondrial metabolism. Although SIRT3 catalyzes the removal of acetyl groups in mitochondria, the addition of the acetyl groups is thought to be primarily controlled in an enzyme-independent manner due to the absence of potent acetyltransferases. In this study, we developed an engineered mitochondria-localized acetyltransferase, named engineered mitochondrial acetyltransferase (eMAT). eMAT localized in the mitochondrial matrix and introduced robust global protein lysine acetylation, including 413 proteins with 1,119 target lysine residues. Notably, 74% of the acetylated proteins overlapped with previously known acetylated proteins, indicating that the eMAT-mediated acetylation system is physiologically relevant. Functionally, eMAT negatively regulated mitochondrial energy metabolism, inhibited cell growth, and promoted cellular senescence, suggesting that mitochondrial hyper-acetylation drives metabolic inhibition and cellular senescence. SIRT3 counteracted eMAT-induced acetylation and metabolic inhibition, restored cell growth, and protected cells from senescence, highlighting the contribution of SIRT3 in maintaining energy metabolism and preventing cellular senescence.