1 GCN 2 is required to increase fibroblast growth factor 21 and maintain hepatic 1 triglyceride homeostasis during asparaginase treatment

53 The anti-leukemic agent asparaginase triggers the amino acid response (AAR) in liver by 54 activating the eukaryotic initiation factor 2 (eIF2) kinase GCN2. To explore the 55 mechanism by which AAR induction is necessary to mitigate hepatic lipid accumulation 56 and prevent liver dysfunction during continued asparaginase treatment, wild-type and 57 Gcn2 null mice were injected once daily with asparaginase or phosphate buffered saline 58 for up to 14 d. Asparaginase induced mRNA expression of multiple AAR genes and 59 greatly increased circulating concentrations of the metabolic hormone fibroblast growth 60 factor 21 (FGF21) independent of food intake. Loss of GCN2 precluded mRNA 61 expression and circulating levels of FGF21 and blocked mRNA expression of multiple 62 genes regulating lipid synthesis and metabolism including Fas, Ppara, Pparg, Acadm 63 and Scd1 in both liver and white adipose tissue. Furthermore, rates of triglyceride export 64 and protein expression of ApoB 100 were significantly reduced in the livers of Gcn2 null 65 mice treated with asparaginase, providing a mechanistic basis for the increase in hepatic 66 lipid content. Loss of AAR-regulated antioxidant defenses in Gcn2 null livers were 67 signified by reduced Gpx1 gene expression alongside increased lipid peroxidation. 68 Substantial reductions in antithrombin III hepatic expression and activity in the blood of 69 asparaginase-treated Gcn2 null mice indicated liver dysfunction. These results suggest 70 that the ability of the liver to adapt to prolonged asparaginase treatment is influenced by 71 GCN2-directed regulation of FGF21 and oxidative defenses, that when lost, corresponds 72 with maladaptive effects on lipid metabolism and hemostasis. 73 74