Abstract 18: APOBEC1 Complementation Factor: A Novel Regulator of Triglyceride Metabolism

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A recent human rare-variant study on plasma lipids has identified a missense mutation in the A1CF (APOBEC1 complementation factor) gene that significantly associates with TG levels. Expressed in the liver and small intestine, A1CF encodes an RNA binding protein that facilitates APOBEC1’s editing of APOB mRNA, introducing a premature stop codon that yields apoB-48. Surprisingly, despite the importance of apoB-48 in plasma lipid metabolism, prior studies have shown that APOBEC1 deficiency itself does not alter plasma TG levels. Therefore, we hypothesize that A1CF regulates TG metabolism independently of its role in APOB mRNA editing. To evaluate this hypothesis, we used CRISPR-Cas9 to generate whole-body A1cf knockout mice ( A1cf - / - ), knock-in mice homozygous for the TG-associated Gly398Ser mutation ( A1cf GS/GS ), and A1cf - / - rat hepatoma cell lines. Both A1cf GS/GS and A1cf - / - mice had significantly increased fasting plasma TG compared to controls (1.71-fold, N = 22-27, P = 1.5 х 10 -3 ; and 1.67-fold, N = 15-27, P = 1.8 х 10 -3 respectively), recapitulating the human TG phenotype and demonstrating that A1CF loss of function leads to hyperTG. Supporting our hypothesis that this occurs independently of APOBEC1 function, apoB-48 was detected by immunoblot in the plasma and small intestines of A1cf - / - and A1cf GS/GS mice without a significant shift in apoB-100/apo-B48. TG clearance was not decreased in A1cf - / - mice during an oral fat tolerance test (N = 12), but A1CF deficiency did result in increased TG secretion in vivo (1.34-fold, N = 12, P < 0.001), consistent with an observed increase in VLDL-apoB secretion by A1cf - / - hepatoma cells (1.55-fold, P < 0.001). We performed RNA-seq of A1cf - / - and wild-type mouse livers, and while differential expression analysis did not yield candidate mechanisms to explain the TG phenotype, we detected 167 alternative splicing (AS) events (delta PSI > 10). We validated several AS events for metabolism genes such as Khk and Hmgcl by RT-PCR, identifying A1CF’s previously unknown role as a regulator of AS events with clear metabolic relevance. In summary, we have demonstrated in our functional follow-up of a human rare-variant study that A1CF modulates TG metabolism through entirely novel mechanisms independent of APOB mRNA editing.