The A20 binding inhibitor of nuclear factor-kappa B (NF-κB)-1 (ABIN-1) serves as a ubiquitin sensor and autophagy receptor, crucial for modulating inflammation and cell death. Our previous in vitro investigation identified the microtubule-associated protein 1A/1B-light chain 3 (LC3)-interacting region (LIR) motifs 1 and 2 of ABIN-1 as key mitophagy regulators. This study aimed to explore the in vivo biological significance of ABIN1-LIR domains using a novel CRISPR-engineered ABIN1-ΔLIR1/2 mouse model, which lacks both the LIR motifs. Comprehensive morphological, serum, and tissue histochemical analyses revealed increased body fat and liver weights, altered serum and hepatic lipid profiles, and substantial hepatic lipid droplet accumulation, indicative of altered hepatic lipid metabolism, dyslipidemia, and hepatic steatosis in ABIN1-ΔLIR1/2 mice. Transcriptomic, metabolomic, and lipidomic analyses indicated dysregulated hepatic mitochondrial metabolism, favoring lipogenesis. Mechanistically, LIR1/2 deletion inhibited the expression and activity of transcription factor EB (TFEB) and AMP-activated protein kinase β1 (AMPKβ1), resulting in compromised autophagy and lipophagy. ABIN1 interacted with TFEB and colocalization was observed in both the cytoplasmic and nuclear compartments of hepatocytes. Impaired mitophagy was evidenced by the decreased expression of parkin and optineurin, along with increased levels of mitochondrial cytochrome c oxidase subunit II. These findings were corroborated by liver biopsies of patients with metabolic dysfunction-associated steatotic liver disease. Thus, this study underscores the functional role of ABIN1-LIR motifs in modulating the ABIN1-AMPK-TFEB axis, which is critical for mitochondria-associated lipid metabolism and mitophagy, offering insights into the mechanistic pathways contributing to the pathogenesis of steatosis-associated liver diseases with potential therapeutic implications.Having identified LC3-interacting region (LIR) motifs 1/2 of A20 binding inhibitor of NF-κB-1 (ABIN-1) as mitophagy regulators in vitro, this study generated CRISPR-engineered ABIN1-ΔLIR1/2 mice lacking both LIR motifs to elucidate its in vivo significance. These mice exhibit enhanced hepatic lipid droplet accumulation, dysregulated mitochondrial metabolism, and impaired mitophagy, through modulation of the AMPK-TFEB axis. Liver biopsies from patients with metabolic dysfunction-associated steatotic liver disease corroborate these findings, suggesting therapeutic implications for liver diseases. NEW & NOTEWORTHY
