Background: Group VIA calcium-independent phospholipase A2 (iPla2 beta) regulates homeostasis and remodeling of phospholipids (PL). We previously showed that iPla2 beta(-/-) mice fed with a methionine-choline-deficient diet (MCD) exhibited exaggerated liver fibrosis. As iPla2 beta is located in the endoplasmic reticulum (ER), we investigated the mechanisms for this by focusing on hepatic ER unfolded protein response (UPR), ER PL, and enterohepatic bile acids (BA). Methods: Female WT (wild-type) and iPla2 beta(-/-) mice were fed with chow or MCD for 5 weeks. PL and BA profiles were measured by liquid chromatography-mass spectrometry. Gene expression analyses were performed. Results: MCD feeding of WT mice caused a decrease of ER PL subclasses, which were further decreased by iPla2 beta deficiency. This deficiency alone or combined with MCD downregulated the expression of liver ER UPR proteins and farnesoid X-activated receptor. The downregulation under MCD was concomitant with an elevation of BA in the liver and peripheral blood and an increase of biliary epithelial cell proliferation measured by cytokeratin 19. Conclusion: iPla2 beta deficiency combined with MCD severely disturbed ER PL composition and caused inactivation of UPR, leading to downregulated Fxr, exacerbated BA, and ductular proliferation. Our study provides insights into iPla2 beta inactivation for injury susceptibility under normal conditions and liver fibrosis and cholangiopathies during MCD feeding.