| NF-κB plays important roles in responsible for extracellular stress stimuli. Extracellular signals are transmitted from membrane receptors to multiple signaling cascades. The stimuli trigger IKK activation and then lead to degradation of IκB proteins, which result in the release of NF-kB proteins from IκB. Then released homo- or hetero- dimmers of NF-κB translocate to nucleus and regulate the transcription of diverse target genes. The regulation of the NF-κB pathway mainly happens on the core components of the pathway including IKKs, IκBs and the NF-κB subunit through multiple posttranslational modi?cations. JNK3, a member of JNK family, can phosphorylate multiple transcription factors such as c-Jun, JunB, JunD, ATF-2 and Elk-1 etc. and some proteins localizing in cytoplasm such as Bcl-2 family proteins, tau and SCG10 etc. Various studies suggest that the JNK signaling pathway is regulated by NF-κB signaling. However, the regulation of NF-κB signaling by JNK pathway remains to be studied.In our previous study, using two yeast-hybrid assay we found that JNK3 interacts with p65 and negatively regulates NF-κB signaling pathway. In the present study we confirm their interaction through co-IP, GST-pulldown and colocalization method. Overexpression of JNK3 can suppress NF-κB transcription activity and co-expression of MKK4 with JNK3 resultin increased interaction between JNK3 and p65 and enhanced the inhibition activity of JNK3 on NF-κB signaling. Using Realtime-PCR analysis, overexpression of JNK3 down-regulates genes associated with adhesion which regulated by NF-κB signaling, and then attenuate cell adhesion ability of hepatocellular carcinoma cell 7402. And we also found that JNK3 can phosphorylate p65 by phosphorylation experiment in vitro.Taken together, we confirmed the interaction between JNK3 and p65 and the negative regulation of NF-κB pathway by JNK3. The phosphoryation of p65 by JNK3 might contribute to the inhibition activity of JNK3 on the NF-κB pathway.
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