The JIP group of mitogen-activated protein kinase scaffold proteins

The response of cells to extracellular stimuli is mediated by evolutionarily conserved signaling mechanisms that regulate proliferation, differentiation, and survival. These mechanisms include mitogen-activated protein kinase (MAPK) signal transduction pathways. Several groups of MAPKs have been identified in yeast, Drosophila melanogaster, and Caenorhabditis elegans. In mammals, three groups of MAPKs have been described: the extracellular signal-regulated protein kinases (ERK), the c-Jun NH2-terminal kinases (JNK), and the p38 MAPKs. Each of these groups of MAPKs is activated by dual phosphorylation on Thr and Tyr by a MAPK kinase (MAPKK) which, in turn, is activated by a MAPKK kinase (MAPKKK). Comparison of ERK, JNK, and p38 MAPKs indicates that the biological function of each group of MAPKs is different. The JNK group of MAPKs is activated by treatment of cells with inflammatory cytokines or by exposure to environmental stress (11). Activated JNK phosphorylates many cellular proteins, including components of the AP-1 transcription factor complex (c-Jun and ATF-2). This phosphorylation leads to increased AP-1 transcription activity (11). The role of increased AP-1 transcription activity is unclear. However, JNK signaling and AP-1 activation have been implicated in multiple biological processes (11). For example, genetic analysis of Drosophila demonstrated that JNK is required for early embryonic morphogenesis (20, 26). Genetic epistasis analysis indicated that the effect of JNK on morphogenesis is mediated by activated Drosophila Jun (9, 19, 25). The JNK signaling pathway is also required for mammalian embryogenesis (6, 14, 17, 34). Three mammalian genes encode JNK protein kinases. JNK1 and JNK2 are expressed ubiquitously, while JNK3 is expressed primarily in the brain (2, 7, 12, 15, 24). A role for JNK in neuronal apoptosis has been demonstrated (31). Recent studies of knockout mice have confirmed this observation. Targeted disruption of the Jnk3 gene causes defects in stress-induced neuronal apoptosis (36), while animals lacking both Jnk1 and Jnk2 genes exhibit defects in developmental neuronal apoptosis (14). JNK is also required for apoptosis of CD4+ CD8+ double-positive thymocytes caused by anti-CD3 in vivo (21, 22). Cellular proliferation, death, and survival may therefore be regulated by the JNK signaling pathway in vivo (11). The JNK signaling pathway also appears to regulate the function of differentiated cells. For example, disruption of the Jnk1 (4) and Jnk2 (22, 35) genes in mice causes defects in T-cell function and immune responses. The JNK signaling pathway therefore contributes to multiple biological processes and represents an important mechanism that is used by cells to respond to extracellular stimulation (11). JNK is activated by phosphorylation on Thr and Tyr by MKK4 and MKK7 (11). These MAPKKs are activated, in turn, by phosphorylation by MAPKKKs, including ASK1, TPL2, TAK1, and members of the MEKK and mixed-lineage protein kinase (MLK) groups of MAPKKKs (11). Biochemical studies demonstrate that each step in the MAPKKK→MAPKK→JNK signaling pathway can be reconstituted in vitro. However, it is unclear whether these assays faithfully mimic the activation of the JNK pathway in vivo. It is likely that the components of the JNK protein kinase cascade may be organized into defined signaling modules (29). For example, the MAPKKK MEKK1 binds to JNK, MKK4, and the Ste20-related protein kinase NIK (27, 30, 32). These interactions may participate in the transmission of signals from MEKK1 to JNK by the creation of a specific signaling module in vivo (29). A functional signaling module could also be created by the interaction of components of the JNK signaling pathway with other proteins. An example is provided by the scaffold protein JNK-interacting protein 1 (JIP1) (3), which binds JNK, MKK7, MLKs, and the Ste20-related protein kinase HPK1 (28). The JIP1 scaffold mediates signaling to JNK by members of the MLK group of MAPKKKs but does not participate in signaling by the MEKK group of MAPKKKs (28). The purpose of the study described in this report was to examine the JIP-mediated JNK signaling module. We demonstrate that JIP1 is a member of a group of MAPK scaffold proteins that includes JIP2. Both JIP1 and JIP2 form homo- and hetero-oligomeric complexes with components of the JNK signaling pathway. The JIP scaffolds facilitate JNK activation by MLK protein kinases by aggregating components of the MAPK cascade to form a functional JNK signaling module.