Abstract

Background: Liver acetylome is a set of protein acetylation’s whose level reflects cellular metabolic health and is directly linked to intracellular pathways. However, to date, little is known about the cellular pathways that maintain the hepatic acetylome levels. Here, we show that macroautophagy hereafter referred to as autophagy, an intracellular lysosomal degradative pathway, regulates the hepatic acetylome.

Methods: Various mouse models of autophagy deficient and autophagy activated conditions were used to dissect the role of autophagy in hepatic acetylome regulation. Wild type (WT) mice were injected with chloroquine (60mg/Kg, i.p) for 6 consecutive days to inhibit autophagy. Total liver lysate and subcellular fractions were analyzed by immunoblotting for acetylated lysine. Liver sections were immunostained for acetyl-Lysine. Hepatic level of acetyl-CoA, CoA and Histone H3 acetylation, and quantitative PCR of various enzymes involved in acetyl-CoA regulation were examined. For rescue experiments, acetyl-CoA was administered intraperitoneally (10mg/kg body weight) for eight consecutive days. Liver sections were subjected to H&E staining and serum ALT was examined as a measure of liver injury.

Results: Examination of total acetylome (nuclear, cytosolic, mitochondrial, membrane) in autophagy-deficient or autophagy-defective liver exhibited remarkably lower levels compared to normal liver. The lower hepatic acetylome was independent of the cellular injury that is commonly seen in autophagy-deficient conditions. In contrast, autophagy activation by fasting or rapamycin treatment increased the level of hepatic acetylome. Moreover, mechanistic studies showed that hepatic autophagy function is essential to maintaining levels of acetyl-CoA, a central intermediate metabolite needed for acetylation of proteins. Autophagy impairment significantly reduced hepatic acetyl-CoA production through transcriptional downregulation of key enzymes involved in the acetyl-CoA synthesis, including Acly, AceCS1, AceCS2, Mcd, and Pdha1. Notably, replenishing hepatic acetyl-CoA rescued the lowered hepatic acetylome and, interestingly, protected against liver injury in the autophagy-deficient liver.

Conclusion: In conclusion, autophagy regulates the hepatic acetylome as an important mechanism for protecting liver against injury and causing liver damage.