Figure 1.

nTrip6 Is a Fos Selective Coactivator

A, nTrip6 specifically interacts in vitro with Fos family members. ³⁵S-labeled, in vitro-translated Fos family members (c-Fos, Fra1, and 2, FosB), Jun family members (c-Jun, JunB, and JunD), or ATF2 were subjected to GST pull-down assays using either GST or GST-Trip6^190–476. Input represents 10% of the in vitro-translated material used in each assay. B, nTrip6 does not interact with c-Jun:ATF2 in living cells. Cos7 cells were transfected with either the single-chain AP-1 c-Jun∼c-Fos fused to YC and Trip6^190–476 fused to YN or the single-chain AP-1 c-Jun∼ATF2 fused to YC and Trip6^190–476 fused to YN, together with a mCherry expression vector as a transfection control. Nuclei were counterstained with DRAQ5. The reconstituted YFP fluorescence was allowed to mature for 1 h at 30 C before live cell imaging. The YFP complementation was observed in 80–90% of the cells transfected with c-Jun∼c-Fos-YC and Trip6^190–476-YN. C, NIH-3T3 cells were transfected with either a control siRNA (Con) or a siRNA targeting Trip6, together with the −1977/−1858uPA-TATA-luciferase reporter gene and Ubi-Renilla. Cells were induced with either TPA or UV 48 h after transfection, and luciferase activities were determined 16 h later. Top, Luciferase activities normalized to Renilla activities are plotted as fold induction (mean ± sd of one representative experiment performed in triplicate); bottom, cell lysates subjected to Western blotting using anti-Trip6, anti-c-Fos, and anti-c-Jun antibodies.