The p38 mitogen-activated protein kinase (MAPK) signaling pathway, acting through the downstream kinase MK2, regulates the stability of many proinflammatory mRNAs that contain adenosine/uridine-rich elements (AREs). It is thought to do this by modulating the expression or activity of ARE-binding proteins that regulate mRNA turnover. MK2 phosphorylates the ARE-binding and mRNA-destabilizing protein tristetraprolin (TTP) at serines 52 and 178. Here we show that the p38 MAPK pathway regulates the subcellular localization and stability of TTP protein. A p38 MAPK inhibitor causes rapid dephosphorylation of TTP, relocalization from the cytoplasm to the nucleus, and degradation by the 20S/26S proteasome. Hence, continuous activity of the p38 MAPK pathway is required to maintain the phosphorylation status, cytoplasmic localization, and stability of TTP protein. The regulation of both subcellular localization and protein stability is dependent on MK2 and on the integrity of serines 52 and 178. Furthermore, the extracellular signal-regulated kinase (ERK) pathway synergizes with the p38 MAPK pathway to regulate both stability and localization of TTP. This effect is independent of kinases that are known to be synergistically activated by ERK and p38 MAPK. We present a model for the actions of TTP and the p38 MAPK pathway during distinct phases of the inflammatory response.The tandem zinc finger protein tristetraprolin (TTP; also known as Nup475, Tis11, or Zfp36) (23,26,40,46,62) is expressed in activated monocytic cells (13, 47) and T lymphocytes (49, 51). It functions to regulate the expression of tumor necrosis factor ␣ (TNF-␣) by binding to a conserved adenosine/uridine-rich element (ARE) within the 3Ј-untranslated region of TNF-␣ mRNA (13,31,32,36,47). TTP promotes both mRNA deadenylation and 3Ј to 5Ј degradation of the mRNA body (35, 37-39), consistent with its ability to recruit several factors involved in these processes (14,25,39,45). The pivotal role of TTP in the regulation of TNF-␣ is illustrated by the proinflammatory phenotype of a TTP Ϫ/Ϫ mouse strain, in which chronic overexpression of TNF-␣ by macrophages results in severe polyarthritis and cachexia (11,13,57). TTP has also been implicated in the posttranscriptional regulation of granulocyte-macrophage colony-stimulating factor (12), interleukin-2 (51), cyclooxygenase 2 (COX-2) (50), and inducible nitric oxide synthase (24). It may also regulate its own expression by binding to an ARE in the 3Ј untranslated region of TTP mRNA (60). The minimum binding site of TTP is the nonameric sequence UUAUUUAUU (2,3,38,65), and it is likely that additional posttranscriptional targets of TTP containing this sequence remain to be identified.The p38 mitogen-activated protein kinase (MAPK) and its downstream kinase MK2 play a central role in the posttranscriptional regulation of inflammatory gene expression in myeloid and other cells (5, 16, 20-22, 33, 34, 54). We and others have therefore investigated interactions of the p38 MAPK pathway with TTP. In a mouse macrophage-like...