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. 2006 Oct 9;175(1):121–133. doi: 10.1083/jcb.200604129

Figure 6.

Figure 6.

Reduced JNK activation in response to TNFα stimulation in FIP200 KO cells and embryos. (A–C) FIP200 KO and WT MEFs were serum starved overnight. They were then left untreated or treated with 50 ng/ml TNFα for the different periods of time, as indicated. Cell lysates were then analyzed by Western blotting with various antibodies as indicated. (D) FIP200 KO and WT MEFs were infected with recombinant adenoviruses encoding FIP200 (Ad-FIP200) or GFP (Ad-GFP, as control), as indicated. 1 d after infection, cells were serum starved overnight and were left untreated or treated with 50 ng/ml TNFα for 10 min. Cell lysates were analyzed by Western blotting with various antibodies as indicated. (E and F) FIP200 KO MEFs were transfected with empty vector or vector encoding Myc-JNK1, along with pEGFP (3:1 ratio). The cells were either left untreated or treated for 1 d with 50 ng/ml TNFα, as indicated. Cell lysates were prepared and analyzed by Western blotting with various antibodies, as indicated (E). Alternatively, the cells were stained with Hoechst to determine the fraction of apoptotic cells in EGFP-positive populations. The mean + SEM from at least three experiments is shown (F). (G) Western blotting analysis of heart and liver protein extracts from FIP200 WT (+/+) and KO (Δ/Δ) embryos using antibodies against phospho-JNK or JNK, as indicated. (H) Hepatocytes isolated from FIP200 WT (+/+) and KO (Δ/Δ) embryos were serum starved overnight. They were then left untreated or treated with 50 ng/ml TNFα for the different periods of time, as indicated. Cell lysates were then analyzed by Western blotting with phospho-JNK or JNK antibodies, as indicated. (I) FIP200 KO hepatocytes were transfected with empty vector or vector encoding Myc-JNK1, along with pEGFP (3:1 ratio). The cells were either left untreated or treated for 1 d with 50 ng/ml TNFα, as indicated. They were then stained with Hoechst to determine the fraction of apoptotic cells in EGFP-positive populations. The mean + SEM from at least three experiments is shown.