Signal Transduction Cross Talk Mediated by Jun N-Terminal Kinase-Interacting Protein and Insulin Receptor Substrate Scaffold Protein Complexes

Scaffold proteins have been established as important mediators of signal transduction specificity. The insulin receptor substrate (IRS) proteins represent a critical group of scaffold proteins that are required for signal transduction by the insulin receptor, including the activation of phosphatidylinositol 3 kinase. The c-Jun NH₂-terminal kinase (JNK)-interacting proteins (JIPs) represent a different group of scaffold molecules that are implicated in the regulation of the JNK. These two signaling pathways are functionally linked because JNK can phosphorylate IRS1 on the negative regulatory site Ser-307. Here we demonstrate the physical association of these signaling pathways using a proteomic approach that identified insulin-regulated complexes of JIPs together with IRS scaffold proteins. Studies using mice with JIP scaffold protein defects confirm that the JIP1 and JIP2 proteins are required for normal glucose homeostasis. Together, these observations demonstrate that JIP proteins can influence insulin-stimulated signal transduction mediated by IRS proteins.The c-Jun NH₂-terminal kinase (JNK)-interacting proteins (JIPs) are implicated in the regulation of the JNK signal transduction pathway (8, 28). The JIP1 and JIP2 proteins are structurally related with similar modular domains (SH3 and PTB) and binding sites for the mixed-lineage protein kinase (MLK) group of mitogen-activated protein kinase (MAPK) kinase kinases, the MAPK kinase MKK7, and JNK (19). These JIP proteins also interact with the microtubule motor protein kinesin, several guanine nucleotide exchange factors, the phosphatase MKP7, Src-related protein kinases, and AKT to form multifunctional protein complexes (19).One potential physiological role of JIP scaffold proteins is the response to metabolic stress, insulin resistance, and diabetes. Several lines of evidence support this hypothesis. First, JIP1 is required for metabolic stress-induced activation of JNK in white adipose tissue (12). Second, MLKs that interact with JIP proteins are implicated as essential components of a signaling pathway that mediates the effects of metabolic stress on JNK activation (13). Third, studies have demonstrated that the human Jip1 gene may contribute to the development of type 2 diabetes, because a Jip1 missense mutation was found to segregate with type 2 diabetes (26). Collectively, these data suggest that JIP proteins play a role in the cellular response to metabolic stress and the regulation of insulin resistance.It is established that the insulin receptor substrate (IRS) group of scaffold proteins plays a central role in insulin signaling (27). Treatment of cells with insulin causes tyrosine phosphorylation of the insulin receptor, the recruitment of IRS proteins to the insulin receptor, and the subsequent tyrosine phosphorylation of IRS proteins on multiple residues that act as docking sites for insulin-regulated signaling molecules, including phosphatidylinositol 3 kinase (27). Negative regulation of IRS proteins is implicated as a mechanism of insulin resistance and can be mediated by multiple pathways, including IRS protein phosphorylation and degradation. Thus, the mTOR/p70^S6K (21, 22, 24) and the SOCS-1/3 (20) signaling pathways can regulate IRS protein degradation. Multisite phosphorylation on Ser/Thr residues can also regulate IRS protein function, including JNK phosphorylation of IRS1 on the inhibitory site Ser-307 that prevents recruitment of IRS1 to the activated insulin receptor (2).The IRS and JIP groups of scaffold proteins may function independently to regulate JNK-dependent and insulin-dependent signal transduction. However, functional connections between these scaffold proteins have been identified. Thus, studies using Jip1^−/− mice demonstrate that JIP1 is required for high-fat-diet-induced JNK activation in white adipose tissue, IRS1 phosphorylation on the inhibitory site Ser-307, and insulin resistance (12). These data suggest that JIP scaffold proteins function cooperatively with IRS proteins to regulate signal transduction by the insulin receptor. The purpose of this study was to examine cross talk between the JIP and IRS scaffold complexes. We demonstrate that the JIP and IRS scaffold complexes physically interact in an insulin-dependent manner and confirm that JIP proteins influence normal glucose homeostasis.