Fig. 6: Trib1 interacts with Nrf2 to block its nuclear translocation.

A Primary murine hepatocytes were transfected with indicated siRNAs followed by treatment with HGF (20 ng/ml) and GW3965 (2 μM) for 24 h. Immunofluorescence staining was performed with anti-Nrf2. B Primary murine hepatocytes were transfected with indicated siRNAs followed by treatment with HGF (20 ng/ml) and GW3965 (2 μM) for 24 h. Nrf2 in cytoplasmic and nuclear fractions was examined by western blotting. C Primary murine hepatocytes were transduced with indicated adenovirus followed by treatment with HGF (20 ng/ml) for 24 h. Immunofluorescence staining was performed with anti-Nrf2. D Primary murine hepatocytes were transduced with indicated adenovirus followed by treatment with HGF (20 ng/ml) for 24 h. Nrf2 in cytoplasmic and nuclear fractions was examined by western blotting. E Immunoprecipitation was performed with anti-Trib1, anti-Nrf2, or IgG using lysates extracted from primary murine hepatocytes. F HEK293 cells were transfected with FLAG-Trib1 and HA-tagged Nrf2. Immunoprecipitation was performed with anti-FLAG. G HEK293 cells were transfected with FLAG-Trib1 and HA-tagged Nrf2. Immunoprecipitation was performed with anti-HA. (H) An ARE reporter was transfected into HepG2 cells with wild-type or truncated Trib1 followed by treatment with HGF (20 ng/ml) for 24 h. Luciferase activities were normalized by protein concentration and GFP fluorescence. I–K Primary murine hepatocytes were transduced with indicated adenovirus followed by treatment with HGF (20 ng/ml) for 24 h. I ROS levels. J Hepatic GSH levels. K Gene expression levels were examined by qPCR. N = 3 biological replicates. Data are expressed as mean ± S.D. *p < 0.05, one-way ANOVA with post-hoc Scheff´e.