Mitogen-activated protein kinases (MAPKs) regulate critical signaling pathways involved in cell proliferation, differentiation and apoptosis. Recent studies have shown that a novel class of scaffold proteins mediates the structural and functional organization of the three-tier MAPK module. By linking the MAP3K, MAP2K and MAPK into a multienzyme complex, these MAPKspecific scaffold proteins provide an insulated physical conduit through which signals from the respective MAPK can be transmitted to the appropriate spatiotemporal cellular loci. Scaffold proteins play a determinant role in modulating the signaling strength of their cognate MAPK module by regulating the signal amplitude and duration. The scaffold proteins themselves are finely regulated resulting in dynamic intra-and inter-molecular interactions that can modulate the signaling outputs of MAPK modules. This review focuses on defining the diverse mechanisms by which these scaffold proteins interact with their respective MAPK modules and the role of such interactions in the spatiotemporal organization as well as context-specific signaling of the different MAPK modules.

T he mitogen-activated protein kinase (MAPK) family consists of a group of kinases responsive to a variety of environmental stimuli. MAPKs can be subdivided into three groups: extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38MAPK (1-8). Although ERK has been shown to be activated primarily by proliferative signals, JNK and p38MAPK are activated by genotoxic as well as cytotoxic stress signals (7-10). The structural organization of these kinases into specific signaling modules appears to be facilitated by scaffolding proteins such as STE5 in yeast (11)(12)(13)(14) and JNK͞stress-activated protein kinase-associated protein (JSAP) and JNK interacting proteins 1-3 in mammalian cells (15)(16)(17)(18)(19)(20). These scaffolding proteins tether different MAPK kinase kinases (MEKKs), MAPK kinases (MKKs), and MAPKs into close proximity so that the successive phosphorylation events can occur efficiently, thus conferring specificity to a particular combination of kinases for activation. Although it is well documented that these phosphorylation cascades lead to the activation of transcription factors such as Fos, Jun, and Myc, the precise mechanisms through which the specific transcription factors are recruited is not known (1, 4). In our search for proteins that associate with transcription factors such as Max and Myc, we identified a scaffolding protein, which we termed JLP for JNK-associated leucine zipper protein. Here we show that JLP brings together Max and c-Myc along with JNK and p38MAPK, as well as their upstream kinases MKK4 and MEKK3. Thus, JLP defines a family of scaffolding proteins that bring MAPKs and their target transcription factors together for the execution of specific signaling pathways. Materials and MethodsCloning of JLP. Human Max was expressed as bacterial recombinant protein, which was 32 P-labeled in vitro with heart muscle kinase (Sigma) by using [␥-32 P]ATP. The Max probe was added in the hybridization buffer Hyb75 (20 mM Hepes-KOH, pH 7.7͞75 mM KCl͞0.1 mM EDTA͞2.5 mM MgCl 2 ͞0.05% Nonidet P-40͞1% nonfat milk͞10 mM DTT) and used to screen the gt11 expression library derived from 32Dcl3 cells (21) as described (22). Four overlapping cDNAs were used to generate a cDNA sequence encoding for the full-length protein of JLP.S Tagging and Mutation of JLP. PCR was used to generate a fragment of JLP sequence (3,250-4,083 bp) whose 3Ј end contained the coding sequence of S tag (KETAAAKFERQH-MDS) followed by a stop codon. Substitution of all leucine residues [amino acids 117, 124, 131, 145, 152, and 159 in leucine zipper I (LZI) and amino acids 413 and 420 in leucine zipper II (LZII)] of JLP with alanine residues was carried out by sitedirected mutagenesis and fusion PCR by using M2 cDNA as the template as described (23). All mutations and deletion constructs were verified by sequence analysis. JLP-S 3Ј deletion mutants were created by digestion of the WT JLP-S cDNA EcoRI͞MluI fragment with SmlI, MslI, PvuII, BssSI, SpeI, and Bsu36I. JLP-S domain constructs were generate...

The p38 mitogen-activated protein kinase (MAPK) pathway plays an important role in cell differentiation, but the signaling mechanisms by which it is activated during this process are largely unknown. Cdo is an immunoglobulin superfamily member that functions as a component of multiprotein cell surface complexes to promote myogenesis. In this study, we report that the Cdo intracellular region interacts with JLP, a scaffold protein for the p38α/β MAPK pathway. Cdo, JLP, and p38α/β form complexes in differentiating myoblasts, and Cdo and JLP cooperate to enhance levels of active p38α/β in transfectants. Primary myoblasts from Cdo −/− mice, which display a defective differentiation program, are deficient in p38α/β activity, and the expression of an activated form of MKK6 (an immediate upstream activator of p38) rescues the ability of Cdo −/− cells to differentiate. These results document a novel mechanism of signaling during cell differentiation: the interaction of a MAPK scaffold protein with a cell surface receptor.

No abstract

In a group of patients with low DO(2)I following major abdominal surgery, microvascular flow abnormalities were more frequent in patients who developed complications. However, there were no differences in DO(2)I, cutaneous PtO(2) or red cell flux between the two groups. Impaired microvascular flow may be associated with the development of post-operative complications.