Transcriptional regulation of tristetraprolin by NF-κB signaling in LPS-stimulated macrophages

Lipopolysaccharide (LPS) treatment causes the marked changes of gene expression in macrophages. Tristetraprolin (TTP), which is an mRNA-destabilizing protein that negatively regulates the expression of pro-inflammatory mediators, is induced by LPS. To delineate the molecular mechanism of LPS-stimulated TTP expression, several inhibitors blocking different signaling pathways were used initially. We observed that inhibitors of the NF-kappa B signaling pathway could down-regulate the TTP expression during LPS-induction. Consistently, TTP expression was increased upon recombinant TNF alpha stimulation which activates NF-kappa B signaling. The 5' regulatory region of zfp36 gene spanning from -2 k to +50 was isolated, which contained a putative NF-kappa B-binding site located in -1859 to -1850. Analysis of luciferase reporter activity driven by a serial 5'-deletion of TTP promoter showed that NF-kappa B inhibitor-mediated suppression of LPS or TNF alpha induced activity was through the predicted kappa B-binding sites. When the NF-kappa B-binding site was mutated, the TTP promoter decreased its response to the ectopic expression of NF-kappa B. Physical interaction analysis including oligonucleotides competition, gel shift and chromatin immunoprecipitation assays demonstrated that NF-kappa B activated by LPS or TNF alpha bound to the TTP promoter specifically. These results suggested that during LPS stimulation, NF-kappa B signaling were activated to regulate the transcription of TTP mRNA.