Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Raf-1 is a serine/threonine kinase which is essential in cell growth and differentiation. Tyrosine kinase oncogenes and receptors and p21ras can activate Raf-1, and recent studies have suggested that Raf-1 functions upstream of MEK (MAP/ERK kinase), which phosphorylates and activates ERK. To determine whether or not Raf-1 directly activates MEK, we developed an in vitro assay with purified recombinant proteins. Epitope-tagged versions of Raf-1 and MEK and kinase-inactive mutants of each protein were expressed in Sf9 cells, and ERK1 was purified as a glutathione S-transferase fusion protein from bacteria. Raf-1 purified from Sf9 cells which had been coinfected with v-src or v-ras was able to phosphorylate kinase-active and kinase-inactive MEK. A kinase-inactive version of Raf-1 purified from cells that had been coinfected with v-src or v-ras was not able to phosphorylate MEK. Raf-1 phosphorylation of MEK activated it, as judged by its ability to stimulate the phosphorylation of myelin basic protein by glutathione S-transferase-ERK1. We conclude that MEK is a direct substrate of Raf-1 and that the activation of MEK by Raf-1 is due to phosphorylation by Raf-1, which is sufficient for MEK activation. We also tested the ability of protein kinase C to activate Raf-1 and found that, although protein kinase C phosphorylation of Raf-1 was able to stimulate its autokinase activity, it did not stimulate its ability to phosphorylate MEK.
Mol Cell Biol 1993 Nov
PMID:Reconstitution of the Raf-1-MEK-ERK signal transduction pathway in vitro. 841 57

We have previously shown that progesterone, but not estradiol or testosterone, can compete with [3H]N-methyl scopolamine (NMS) for the cardiac M2 muscarinic binding site. Experiments have been carried out to investigate the inhibitory effects of a large variety of progesterone-like steroids on the M2 muscarinic receptor. These studies were performed with the aid of a new binding assay which uses intact tissue in the form of ventricular micropunches. Our data show that synthetic, clinically-used progestins such as Provera, norgesterel and cyproterone are largely ineffective at the M2 binding site whereas the naturally occurring progesterone derivatives 17 alpha hydroxy progesterone and Reichstein's Substance S are highly active. (Ki values 5 x 10(-6)M and 1 x 10(-6)M, respectively). Minor structural modifications such as acetylation of the 17 alpha hydroxy group abolishes activity. Steroids known to exert cell membrane effects, such as alfaxalone and pregnenolone sulfate, had no influence on [3H] NMS binding. The progesterone antagonist RU-486 did not block the inhibitory effect of progesterone. Moreover, this putative receptor may be located in the cardiomyocyte membrane since 17 alpha hydroxy progesterone induces rapid dissociation of [3H] NMS from its binding site (30% reduction in 5 min). We attempted to further localize the inhibitory locus to the M2 receptor itself by means of the irreversible antagonist propylbenzilylcholine mustard (PrBCM). Ninety percent of the M2 receptor could be blocked by PrBCM (10(-6) M), an effect reversible by the specific muscarinic antagonist scopolamine methyl bromide but not by progesterone. These results suggested that progesterone does not interact directly with the [3H] NMS labelled M2 binding site.(ABSTRACT TRUNCATED AT 250 WORDS)
J Mol Cell Cardiol 1995 Sep
PMID:Modulation of cardiac M2 muscarinic receptor binding by progesterone-related steroids. 852 44

The HST7 gene of Candida albicans encodes a protein with structural similarity to MAP kinase kinases. Expression of this gene in Saccharomyces cerevisiae complements disruption of the Ste7 MAP kinase kinase required for both mating in haploid cells and pseudohyphal growth in diploids. However, Hst7 expression does not complement loss of either the Pbs2 (Hog4) MAP kinase kinase required for response to high osmolarity, or loss of the Mkk1 and Mkk2 MAP kinase kinases required for proper cell wall biosynthesis. Intriguingly, HST7 acts as a hyperactive allele of STE7; expression of Hst7 activates the mating pathway even in the absence of upstream signaling components including the Ste7 regulator Ste11, elevates the basal level of the pheromone-inducible FUS1 gene, and amplifies the pseudohyphal growth response in diploid cells. Thus Hst7 appears to be at least partially independent of upstream activators or regulators, but selective in its activity on downstream target MAP kinases. Creation of Hst7/Ste7 hybrid proteins revealed that the C-terminal two-thirds of Hst7, which contains the protein kinase domain, is sufficient to confer this partial independence of upstream activators.
Mol Gen Genet 1995 Dec 20
PMID:Constitutive activation of the Saccharomyces cerevisiae mating response pathway by a MAP kinase kinase from Candida albicans. 854 26

Tyrosine kinase growth factor receptors activate MAP kinase by a complex mechanism involving the SH2/3 protein Grb2, the exchange protein Sos, and Ras. The GTP-bound Ras protein binds to the Raf kinase and initiates a protein kinase cascade that leads to MAP kinase activation. Three MAP kinase kinase kinases have been described--c-Raf, c-Mos, and Mekk--that phosphorylate and activate Mek, the MAP kinase kinase. Activated Mek phosphorylates and activates MAP kinase. Subsequently, the activated MAP kinase translocates into the nucleus where many of the physiological targets of the MAP kinase signal transduction pathway are located. These substrates include transcription factors that are regulated by MAP kinase phosphorylation (e.g., Elk-1, c-Myc, c-Jun, c-Fos, and C/EBP beta). Thus the MAP kinase pathway represents a significant mechanism of signal transduction by growth factor receptors from the cell surface to the nucleus that results in the regulation of gene expression. Three MAP kinase homologs have been identified in the rat: Erk1, Erk2, and Erk3. Human MAP kinases that are similar to the rat Erk kinases have also been identified by molecular cloning. The human Erk1 protein kinase has been shown to be widely expressed as a 44-kDa protein in many tissues. The human Erk2 protein kinase is a 41-kDa protein that is expressed ubiquitously. In contrast, a human Erk3-related protein kinase has been found to be expressed at a high level only in heart muscle and brain. The loci of these MAP kinase genes are widely distributed within the human genome: erk2 at 22q11.2; erk1 at 16p11.2; and ek3-related at 18q12-21. In the yeast Saccharomyces cerevisiae, five MAP kinase gene homologs have been described: smkl, mpk1, hog1, fus3, and kss1. Together, these kinases are a more diverse group than the human erks that have been identified. Thus the erks are likely to represent only one subgroup of a larger human MAP kinase gene family. A candidate for this extended family of MAP kinases is the c-Jun NH2-terminal kinase (Jnk), which binds to and phosphorylates the transcription factor c-Jun at the activating sites Ser-63 and Ser-73. Evidence is presented here to demonstrate that Jnk is a distant relative of the MAP kinase group that is activated by dual phosphorylation at Tyr and Thr.
Mol Reprod Dev 1995 Dec
PMID:Transcriptional regulation by MAP kinases. 860 77

Members of the Ras superfamily of proteins function as regulated GDP/GTP switches that cycle between active GTP-complexed and inactive GDP-complexed states. Guanine nucleotide exchange factors (GEFs) stimulate formation of the GTP-bound state, whereas GTPase activating proteins (GAPs) catalyze the formation of the GDP-bound state. We describe three studies that evaluate the mechanism of action of GEFs for Ras (SOS1 and RasGRF/CDC25) or Ras-related Rho (Dbl and Vav) proteins. Growth factor-mediated activation of Ras is believed to be mediated by activation of Ras GEFs (CDC25/GRF and SOS1/2). Although the mechanisms of Ras GEF regulation are unclear, recent studies suggest that translocation of SOS1 to the plasma membrane, where Ras is located, might be responsible for Ras activation. Our observation that the addition of the Ras plasma membrane-targeting sequence to the catalytic domains of CDC25 and SOS1 greatly enhanced their transforming and transactivation activities (10-50 fold and 5-10 fold, respectively) suggests that membrane translocation alone is sufficient to potentiate GEF activation of Ras. We have determined that two Ras-related proteins, designated R-Ras and R-Ras2/TC21, can trigger the malignant transformation of NIH 3T3 cells via activation of the Ras signal transduction pathway. Furthermore, like Ras and R-Ras, we observed that TC21 GTPase activity was stimulated by Ras GAPs. However, we observed that both SOS1 and CDC25 were activators of normal TC21, but not R-Ras, transforming activities. Therefore, TC21, but not R-Ras, may be activated by the same extracellular signaling events that activate Ras proteins. Dbl family proteins are believed to function as GEFs and activators of the Ras-related Rho family of proteins. However, one Dbl family oncogene, designated Vav, has been reported to be a GEF for Ras proteins. Therefore we were interested in determining whether Dbl family oncogenes cause transformation by triggering the constitutive activation of Rho or Ras proteins. Our results suggest that Dbl oncogenes cause transformation via a Ras-independent activation of MAP kinases and Rho family proteins.
Mol Reprod Dev 1995 Dec
PMID:Guanine nucleotide exchange factors: activators of Ras superfamily proteins. 860 78

A peptide corresponding to the second complementarity determining region of the heavy chain (CDR2 VH) from a murine anti-CD4 monoclonal antibody, designated L202, was synthesized by solid phase methodology in a number of different antigenic forms, for the purpose of comparing the effectiveness of different adjuvant-carrier systems in the induction of a murine antibody response against the immunizing peptide and parent antibody molecule. Two of the synthetic constructs contained the palmitoyl and N-palmitoyl-cysteinyl-S-(2,3-palmitoyloxy)-propanediol (PAM3Cys) moieties, respectively, attached to the peptide amino terminus with the immunogen comprising liposomal formulations of each. A third immunogen consisted of the CDR2 VH peptide admixed with the PAM3Cys non-covalently and incorporated into liposomes (PAM3Cys + CDR2 VH). A fourth composition comprised the CDR2 VH peptide conjugated to KLH via the sulfhydryl of an added N terminal cysteine (KLH-CDR2 VH) and injected with Complete Freund's adjuvant (CFA). A fifth immunogen consisted of the CDR2 VH peptide synthesized on an octameric, branched polylysine core as a multiple antigenic peptide (MAP-CDR2 VH) injected in the presence of Freund's adjuvant. Groups of five mice were injected intramuscularly with each of these immunogens and bled at two week intervals. The highest anti-peptide gamma-immunoglobulin (IgG) responses (against uncoupled peptide by ELISA) after 56 days were obtained with mice receiving the PAM3Cys-CDR2 VH peptide. However, when screened against the CDR2 V(H) peptide present as the MAP derivative by ELISA, IgG raised against the cognate MAP-CDR2 peptide was much more reactive than IgG raised against the liposomal PAM3Cys-CDR2 VH immunogen. In either case, IgG raised against the KLH-CDR2VH conjugate was poorly reactive. These differences in reactivity to the two forms of the CDR2 VH peptide by ELISA did not correspond to major differences in reactivities to the intact L202 Ab by ELISA. Although the IgG against the MAP immunogen was slightly more reactive than the other antisera against the l202 Ab, all titers were less than 1:100. These data illustrate some limitations of using anti-peptide responses as indicators of potential reactivity against the native protein, but suggest that alternate formulations including lipoidal peptides are more effective than corresponding KLH-peptide conjugates in eliciting Ab responses against poorly immunogenic epitopes.
Mol Immunol 1995 Dec
PMID:Dependence of the murine antibody response to an anti-CDR2 VH peptide on immunogen formulation. 864 1

Microtubule-associated protein-2 (MAP2) is the most abundant MAP in neurons, where its distribution is restricted to the somatodendritic compartment. This molecule undergoes developmentally regulated alternative splicing, resulting in at least two isoforms, a juvenile isoform (termed MAP2c) and a mature isoform (MAP2), with greatly different molecular masses. Spodoptera frugiperda (Sf9) cell expression of the juvenile versus the mature MAP2 isoform generates two distinct patterns of process outgrowth. The smaller juvenile isoform induces multiple short thin processes. Mature MAP2 tends to induce single processes that are considerably thicker than those processes induced by juvenile MAP2. We found important differences in the variability of spacing between microtubules and the number of microtubules along the processes induced by MAP2c and mature MAP2. MAP2c showed variability with most microtubules spaced as closely as with tau, but some spaced as far apart as with mature MAP2. Over their length, the mature MAP2 processes demonstrate proximo-distal taper, which corresponds to a narrowing of the spacing between microtubules from 90 nm to 40 nm. Moreover, there is a decreased number of microtubules in mature MAP2-induced processes whereas in tau and MAP2-induced processes, the number of microtubules is constant along the length. Based on these observations, we conclude that MAP2 isoforms can serve as architectural elements by establishing specific morphological features of processes and specific arrangements of their microtubules.
Mol Biol Cell 1996 Mar
PMID:Juvenile and mature MAP2 isoforms induce distinct patterns of process outgrowth. 886 72

The immediate early gene-encoded enzyme, MAP kinase phosphatase 1 (MKP-1), is thought to be a key element in controlling cellular signalling pathways activated by MAP kinases. Since MAP kinase have been demonstrated to participate in neuronal stimulus-transcription coupling following seizure activity, the present study investigated the induction of MKP-1 in the rat brain after limbic epilepsy. MKP-1 expression was studied with a polyclonal antiserum by Western blots, immunocytochemistry and immuno-electron microscopy at different time periods between 1 and 24 h after kainic acid-induced limbic seizures. MKP-1 induction was identified in dentate granule cells of the hippocampus but not in pyramidal neurons, furthermore in neurons of the outer layers of the neocortex, as well as in neurons of the lateral nucleus of the bed of the stria terminalis. Immuno-electron microscopy demonstrated that MKP-1 was localized in the neuronal nucleus, where the substrate of MKP-1, activated MAP kinases, are also found. In view of the restricted areas of MKP-1 expression and the widespread areas of altered MAP kinases activity it can be concluded that in the majority of CNS populations other mechanisms than MKP-1 induction are responsible for the shut-off of MAP kinases following seizure activity. MKP-1 may contribute in the specific subpopulations where it is induced to the post-translational control of inducible transcription factors of the fos, jun and myc family.
Brain Res Mol Brain Res 1996 Sep 05
PMID:Transient expression of the mitogen-activated protein kinase phosphatase MKP-1 (3CH134/ERP1) in the rat brain after limbic epilepsy. 888 36

Successful axon regeneration requires that signals from the site of injury reach the nucleus to elicit changes in transcription. In spite of their obvious importance, relatively few of these signals have been identified. Recent work on regeneration in the marine mollusk Aplysia californica has provided several insights into the molecular events that occur in neurons after axon injury. Based on these findings, we propose a model in which axon regeneration is viewed as the culmination of a series of temporally distinct but overlapping phases. Within each phase, specific signals enter the nucleus to prime the cell for the arrival of subsequent signals. The first phase begins with the arrival of injury-induced action potentials, which act via calcium and cAMP to turn on genes used in the early stages of repair. In the next phase, MAP-kinases and other intrinsic constituents activated at the injury site are retrogradely transported through the axon to the nucleus, informing the nucleus of the severity of the axonal injury, reinforcing the earlier events, and triggering additional changes. The third phase is characterized by the arrival of signals that originate from extrinsic growth factors and cytokines released by cells at the site of injury. In the last phase, signals from target-derived growth factors arrive in the cell soma to stop growth. Because many of these events appear to be universal, this framework may be useful in studies of nerve repair in both invertebrates and vertebrates.
Mol Neurobiol 1996 Aug
PMID:Priming events and retrograde injury signals. A new perspective on the cellular and molecular biology of nerve regeneration. 889 36

In vivo vanadate and vanadyl have been shown to mimic the action of insulin and to be effective treatment for animal models of both Type I and Type II diabetes. The molecular mechanism of action of the vanadium salts on insulin sensitivity remains uncertain, and several potential sites proposed for the insulin-like effects are reviewed. In human trials, insulin sensitivity improved in patients with NIDDM, as well as in some patients with IDDM after two weeks of treatment with sodium metavanadate. This increase in insulin sensitivity was primarily due to an increase in non-oxidative glucose disposal, whereas oxidative glucose disposal and both basal and insulin stimulated suppression of hepatic glucose output (HGP) were unchanged. Clinically, oral vanadate was associated with a small decrease in insulin requirements in IDDM subjects. Of additional benefit, there was a decrease in total cholesterol levels in both IDDM and NIDDM subjects. Furthermore, there was an increase in the basal activities of MAP and S6 kinases to levels similar to the insulin-stimulated levels in controls, but there was little or no further stimulation with insulin was seen. Further understanding of the mechanism of vanadium action may ultimately be useful in the design of drugs that improve glucose tolerance.
Mol Cell Biochem
PMID:In vivo and in vitro studies of vanadate in human and rodent diabetes mellitus. 892 42


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>