Tristetraprolin regulates TNF TNF-α mRNA stability via a proteasome dependent mechanism involving the combined action of the ERK and p38 pathways

Tumor necrosis factor-alpha (TNF-α) is a central mediator of inflammation. TNF-α expression is regulated by transcriptional and post-transcriptional mechanisms, including mRNA stability and translation. Post-transcriptional control operates through cis-elements in the 3′ Untranslated-Region of the TNF-α mRNA to which trans-acting proteins bind. One of the best characterized trans-acting proteins is Tristetraprolin (TTP), which regulates TNF-α message stability. However, the precise mechanisms controlling TNF-α message stability are unclear, with data supporting a role for the proteasome, the exosome, and the RNA processing-body (P-body), as well as the involvement of the microRNAs. We examined the effect of proteasome inhibition on endogenous TNF-α mRNA stability, TNF-α 3′UTR reporter expression and TTP function in the RAW264.7 cells. These data establish that proteasome inhibition stabilized endogenous TNF-α mRNA, increased TTP protein levels but inhibited TTP mediated TNF-α mRNA decay. Importantly, proteasome inhibition stabilized the TNF-α message to the same degree as LPS stimulation. To further characterize the control of TTP function, we examined the combinatorial effect of p38, ERK and JNK activation on TNF-α post-transcriptional expression and TTP function. These data establish that TTP mediated TNF-α mRNA decay is inhibited by the combined activation of ERK and p38 and not by p38 activation alone. The combined activation of ERK/p38 was sufficient to stabilize endogenous TNF-α mRNA to the same degree as LPS stimulation. Together these data indicate that the proteasome is a critical control point for TTP mediated TNF-α mRNA decay and activation of both ERK and p38 is required to inhibit TTP function and stabilize TNF-α mRNA.