| The c-Jun N-terminal kinase (JNK) is generally thought to play roles in inflammation, proliferation, and apoptosis. Accordingly, its substrates are transcription factors and anti-apoptotic proteins. However, JNK has also been Shown to be required for Drosophila dorsal closure, and MEK kinase1, an upstream kinase in the JNK pathway, has been demonstrated to be essential for cell migration. Both results imply that JNK is important in cell migration. Here we show that JNK1 is required for the rapid movement of both fish keratocytes and rat bladder tumor epithelial cells (NBT-II). Moreover, JNK1 phosphorylates serine 178 on paxillin, a focal adhesion adaptor, both in vitro and in intact cells. NBT-II cells expressing PaxS178A formed focal adhesions and exhibited the limited movement associated with Such contacts in both single cell migration and wound healing assays. On the other hand, cells expressing WT paxillin moved rapidly and retained close contacts as the predominant adhesion. Expression of PaxS178A also inhibited the migration of two other cell lines. Thus, phosphorylation of paxillin by JNK appears essential for maintaining the labile adhesions required for rapid cell migration.; Cell adhesions play an important role in neurite extension. Paxillin, a focal adhesion adaptor protein involved in focal adhesion dynamics, has been demonstrated to be required for neurite outgrowth. However, the molecular mechanism by which paxillin regulates neurite outgrowth is unknown. We Show here that paxillin is phosphorylated by p38mapk in vitro and in nerve growth factor (NGF)-induced PC-12 cells. Ser 85 is identified as one of major phosphorylation sites by phosphopeptide mapping and mass spectrometry. Moreover, expression of the Ser 85 → Ala mutant of paxillin (paxS85A) significantly inhibits NGF-induced neurite extension of PC-12 cells, while expression of wild-type (wt) paxillin does not influence neurite outgrowth. Further experiments indicate that cells expressing paxS85A exhibit small, clustered focal adhesions which are not normally seen in cells expressing wt paxillin. Expression of a p38mapk dominant negative mutant (p38αAF) also enhances focal adhesion formation, whereas a constitutively active MAPK kinase for p38mapk (MKK3bE) causes focal adhesion disassembly. Thus, phosphorylation of Ser 85 on paxillin is involved in NGF-induced neurite extension of PC-12 cells, probably through regulating focal adhesion organization. |
