MCPIP1 Enhances TNF-α-Mediated Apoptosis through Downregulation of the NF-κB/cFLIP Axis

Monocyte chemoattractant protein-1-induced protein 1 (MCPIP1) is rapidly produced under proinflammatory stimuli, thereby feeding back to downregulate excessive inflammation. In this study, we used the stable, inducible expressions of wild-type (WT) MCPIP1 and an MCPIP1-D141N mutant in T-REx-293 cells by means of a tetracycline on (Tet-on) system. We found that WT MCPIP1 but not MCPIP1-D141N mutant expression dramatically increased apoptosis, caspase-3, -7, -8, and -9 activation, and c-Jun N-terminal kinase (JNK) phosphorylation in TNF-alpha-treated cells. The pan-caspase inhibitor, z-VAD-fmk, and the caspase-1 inhibitor, z-YVAD-fmk, but not the JNK inhibitor, SP600125, significantly reversed apoptosis and caspase activation in TNF-alpha/MCPIP1-treated cells. Surprisingly, MCPIP1 itself was also cleaved, and the cleavage was suppressed by treatment with the pan-caspase inhibitor and caspase-1 inhibitor. Moreover, MCPIP1 was found to contain a caspase-1/-4 consensus recognition sequence located in residues 234 similar to 238. As expected, the WT MCPIP1 but not the MCPIP1-D141N mutant suppressed NF-kappa B activation, as evidenced by inhibition of I kappa B kinase (IKK) phosphorylation and I kappa B degradation using Western blotting, IKK activity using in vitro kinase activity, and NF-kappa B translocation to nuclei using an immunofluorescence assay. Interestingly, MCPIP1 also significantly inhibited importin alpha 3 and importin alpha 4 expressions, which are major nuclear transporter receptors for NF-kappa B. Inhibition of NF-kappa B activation further downregulated expression of the caspase-8 inhibitor, cFLIP. In summary, the results suggest that MCPIP1 could enhance the TNF-alpha-induced apoptotic pathway through decreasing NF-kappa B activation and cFLIP expression.

Automated summary

MCPIP1 is rapidly produced under proinflammatory stimuli to downregulate excessive inflammation. In this study, the expression of WT MCPIP1 increased apoptosis and caspase activation in TNF-alpha-treated cells. MCPIP1 also suppressed NF-kappa B activation and cFLIP expression, further enhancing the apoptotic pathway induced by TNF-alpha.