Multiple pathways contribute to nonalcoholic fatty liver disease (NAFLD) in response to high fat diets (HFD). A homolog of mammalian JNK-interacting protein 3 (JIP3), also known as JSAP-1, activates different components in various signaling pathways to modulate cellular processes. The purpose of this study was to examine the role of JIP3 in obesity-related pathologies pathway. Wild-type (WT) C57BL/6 and JIP3-knockout (J1P3(-/-)) mice were randomized to chow or HFD. HFD-fed WT mice increased hepatic JIP3 expression. Mice lacking JIP3 exhibited reduced weight gain, hepatic steatosis, insulin resistance, lipid accumulation, oxidative stress and inflammatory response in mice fed a HFD, which were, importantly, dependent on various signaling pathways. Lipogenesis-linked pathway was inhibited in JIP3(-/-) mice after HFD, while PPAR alpha/gamma were increased. Additionally, JIP3-/- inhibited hepatic oxidative stress, evidenced by down-regulation of total reactive oxygen species (ROS), H2O2, O-2(-), malondialdehyde (MDA), xanthine oxidase (XO), inducible nitric oxide synthase (iNOS), and up-regulation of superoxide dismutase (SOD) and total antioxidant capacity (TAC) in mice after HFD feeding, which might be related to nuclear respiratory factor 2 (Nrf-2) pathway activation. Further, inflammatory response was blocked in JIP3(-/-) mice fed with HFD. The process might be attributed to the suppression of toll-like receptors (TLRs), p-nuclear factor kappa B (NF-kappa B) and p-c-Jun-N-terminal kinase (JNK). Thus, JIP3 absence is associated with decreased lipogenesis, oxidative stress and inflammation, supplying a new target for NAFLD treatment. (C) 2018 Elsevier Inc. All rights reserved.