Abstract
The c-Jun N-terminal kinase (JNK) signaling pathway is a major mediator of stress responses in cells. Similar to other mitogen-activated protein kinases (MAPKs), JNK activity is controlled by a cascade of protein kinases and by protein phosphatases, including dual-specificity MAPK phosphatases. Components of the JNK pathway associate with scaffold proteins that modulate their activities and cellular localization. The JNK-interacting protein-1 (JIP-1) scaffold protein specifically binds JNK, MAPK kinase 7 (MKK7), and members of the mixed lineage kinase (MLK) family, and regulates JNK activation in neurons. In this study we demonstrate that distinct regions within the N termini of MKK7 and the MLK family member dual leucine zipper kinase (DLK) mediate their binding to JIP-1. We have also identified amino acids in JNK required for: (a) binding to JIP-1 and for JIP-1-mediated JNK activation, (b) docking to MAPK kinase 4 (MKK4) and efficient phosphorylation by MKK4, and (c) docking to its substrate c-Jun and efficient c-Jun phosphorylation. None of the amino acids identified were essential for JNK docking to MKK7 or the dual-specificity phosphatase MAPK phosphatase 7 (MKP7). These findings uncover molecular determinants of JIP-1 scaffold complex assembly and demonstrate that there are overlapping, but also distinct, binding determinants within JNK that mediate interactions with scaffold proteins, activators, phosphatases, and substrates.
Highlights
The MAPK1 family of protein kinases are critical mediators of cellular responses to many extracellular signals [1, 2]
Regions of dual leucine zipper kinase (DLK) Required for Binding to JNK-interacting protein-1 (JIP-1), MAPK kinase 7 (MKK7), and for Dimerization—The mixed lineage kinase (MLK) family member DLK associates with the C terminus of the scaffold protein JIP-1 [8]
Jun N-terminal kinase (JNK), MKK7, and MLKs bind to distinct regions of JIP-1 [7, 8]
Summary
The MAPK1 family of protein kinases are critical mediators of cellular responses to many extracellular signals [1, 2]. In the present study we have (i) identified the binding determinants within components of this pathway for docking to each other and to the scaffold protein JIP-1, and (ii) used binding-defective mutants to examine the functional consequences of disrupting these interactions on JNK signaling in cells.
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