SETDB1 enhances starvation-induced lipophagy by inhibiting m⁶A-mediated mRNA decay via DDX5 methylation

Lipophagy, a selective form of macroautophagy/autophagy, degrades lipid droplets (LDs) to provide energy and is implicated in metabolic disorders. The molecular mechanism underlying lipophagy induction remains incompletely understood. This study explored the role of SETDB1 in starvation-induced autophagy and lipophagy. We demonstrate that SETDB1 deficiency exacerbates starvation-induced hepatic lipid accumulation by inhibiting lipophagy. Mechanistically, starvation promotes ATM-mediated phosphorylation of SETDB1, which enhances its interaction with and methylation of the RNA helicase DDX5. In SETDB1-knockout hepatocytes, hypomethylation of DDX5 facilitates the formation of the DDX5-METTL3-METTL14 complex, increasing m⁶A modification of BECN1 and TFEB mRNAs. This modification promoted YTHDF2-mediated decay of these transcripts, thereby inhibiting starvation-induced autophagy and lipophagy. Furthermore, administration of the SETDB1 activator (R, R)-59 significantly enhances lipophagy and attenuates starvation-induced hepatic steatosis. Collectively, our findings reveal a novel pathway in which SETDB1 deficiency drives m⁶A-mediated mRNA degradation to suppress lipophagy, thereby contributing to hepatic steatosis.Abbreviations: AA free: amino acid deprivation; ATG14: autophagy related 14; ATG5: autophagy related 5; ATG7: autophagy related 7; ATM: ATM serine/threonine kinase; Baf A1: bafilomycin A₁; DDX5: DEAD-box helicase 5; FASN: fatty acid synthase; LAMP1: lysosome associated membrane protein 1; LAMP2A: lysosome associated membrane protein 2A; LIPE/HSL: lipase E, hormone sensitive type; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MAP3K5/ASK1: mitogen-activated protein kinase kinase kinase 5; METTL3: methyltransferase 3, N6-adenosine-methyltransferase complex catalytic subunit; METTL14: methyltransferase 14, N6-adenosine-methyltransferase non-catalytic subunit; MGLL/MGL: monoglyceride lipase; OA: oleic acid; OSBPL8/ORP8: oxysterol binding protein like 8; PLIN2: perilipin 2; PNPLA2/ATGL: patatin like domain 2, triacylglycerol lipase; SETDB1: SET domain bifurcated histone lysine methyltransferase 1; TFEB: transcription factor EB; TP53/p53: tumor protein p53; ULK1: unc-51 like autophagy activating kinase 1; YTHDF2: YTH N6-methyladenosine RNA binding protein F2.