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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have investigated the effect of glucose deprivation treatment on the activation of mitogen activated protein kinases (MAPKs) in the drug-sensitive human breast carcinoma cells (MCF-7) and its drug resistant variant (MCF-7/ADR) cells. Western blots and in-gel kinase assays showed that glucose free medium was a strong stimulus for the activation of MAPK in MCF-7/ADR cells. No activation was seen in MCF-7 cells. MAPK was activated within 3 min of being in glucose free medium and it remained activated for over 1 h in MCF-7/ADR cells. After being returned to complete medium, 1 h was required for the MAPK to become deactivated. To investigate whether alternative sources of ATP could inhibit glucose deprivation induced MAPK activation, we added glutamine and glutamate to glucose deprived medium. The addition of glutamine did not reverse glucose deprivation induced MAPK activation in MCF-7/ADR cells. The addition of glutamate, however, decreased the MAPK activation and the length of time of activation. We observed an increase greater than three fold in MEK, Raf, Ras, and PKC activity with glucose deprivation in MCF-7/ADR cells. This suggests that glucose deprivation-induced MAPK activation is mediated through this signal transduction pathway.
Mol Cell Biochem 1997 May
PMID:Differential effect of glucose deprivation on MAPK activation in drug sensitive human breast carcinoma MCF-7 and multidrug resistant MCF-7/ADR cells. 914 15

v-H-ras effector mutants have been assessed for transforming activity and for the ability of the encoded proteins to interact with Raf-1-, B-Raf-, byr2-, ralGDS-, and CDC25-encoded proteins in the yeast two-hybrid system. Transformation was assessed in rat2 cells as well as in a mutant cell line, rv68BUR, that affords a more sensitive transformation assay. Selected mutant Ras proteins were also examined for their ability to interact with an amino-terminal fragment of Raf-1 in vitro. Finally, possible cooperation between different v-H-ras effector mutants and between effector mutants and overexpressed Raf-1 was assessed. Ras transforming activity was shown to correlate best with the ability of the encoded protein to interact with Raf-1. No evidence for cooperation between v-H-ras effector mutants was found. Signaling through the Raf1-MEK-mitogen-activated protein kinase cascade may be the only effector pathway contributing to RAS transformation in these cells.
Mol Cell Biol 1997 Jun
PMID:Interaction of activated Ras with Raf-1 alone may be sufficient for transformation of rat2 cells. 915 3

Endothelin is a small peptide that is a potent bronchoconstrictor, mitogen for airway smooth muscle (ASM), and is believed to be involved in the pathogenesis of asthma. To understand how endothelin stimulates the proliferation of ASM cells in culture, we evaluated the relationship between mitogen activated protein (MAP) kinase activation and cell proliferation. Endothelin is a potent stimulator of the extracellular regulated kinase 2 (ERK2) subgroup of MAP kinases, and ERK2 activation was tightly correlated with the proliferation of rat ASM cells. PD98059, a small molecule inhibitor of MEK (MAP or ERK kinase) was used to establish the role of ERK2 activation in the endothelin-stimulated signal transduction pathway leading to cell proliferation. While PD98059 significantly inhibited the ability of endothelin to activate ERK, the drug did not appear to effect the catalytic activity of an activated MEK mutant, or ERK in vitro. The data suggest that the mechanism of PD98059 inhibition of the ERK2 pathway in ASM cells may involve inhibition of MEK activation. The endothelin signal transduction pathway that culminates in ERK2 activation was dependent on protein kinase C (PKC), since depletion of PKC significantly inhibited the ability of endothelin to activate ERK2. Taken together, the data imply that activation of ERK is a critical endpoint in the endothelin signal transduction pathway since inhibition of this kinase inhibits endothelin-induced ASM cell proliferation.
Am J Respir Cell Mol Biol 1997 May
PMID:Inhibition of ERK activation attenuates endothelin-stimulated airway smooth muscle cell proliferation. 916 Aug 41

The ability of basic helix-loop-helix muscle regulatory factors (MRFs), such as MyoD, to convert nonmuscle cells to a myogenic lineage is regulated by numerous growth factor and oncoprotein signaling pathways. Previous studies have shown that H-Ras 12V inhibits differentiation to a skeletal muscle lineage by disrupting MRF function via a mechanism that is independent of the dimerization, DNA binding, and inherent transcriptional activation properties of the proteins. To investigate the intracellular signaling pathway(s) that mediates the inhibition of MRF-induced myogenesis by oncogenic Ras, we tested two transformation-defective H-Ras 12V effector domain variants for their ability to alter terminal differentiation. H-Ras 12V,35S retains the ability to activate the Raf/MEK/mitogen-activated protein (MAP) kinase cascade, whereas H-Ras 12V,40C is unable to interact directly with Raf-1 yet still influences other signaling intermediates, including Rac and Rho. Expression of each H-Ras 12V variant in C3H10T1/2 cells abrogates MyoD-induced activation of the complete myogenic program, suggesting that MAP kinase-dependent and -independent Ras signaling pathways individually block myogenesis in this model system. However, additional studies with constitutively activated Rac1 and RhoA proteins revealed no negative effects on MyoD-induced myogenesis. Similarly, treatment of Ras-inhibited myoblasts with the MEK1 inhibitor PD98059 revealed that elevated MAP kinase activity is not a significant contributor to the H-Ras 12V effect. These data suggest that an additional Ras pathway, distinct from the well-characterized MAP kinase and Rac/Rho pathways known to be important for the transforming function of activated Ras, is primarily responsible for the inhibition of myogenesis by H-Ras 12V.
Mol Cell Biol 1997 Jul
PMID:Signaling through mitogen-activated protein kinase and Rac/Rho does not duplicate the effects of activated Ras on skeletal myogenesis. 919 90

Activation of the Raf serine/threonine protein kinases is tightly regulated by multiple phosphorylation events. Phosphorylation of either tyrosine 340 or 341 in the catalytic domain of Raf-1 has been previously shown to induce the ability of the protein kinase to phosphorylate MEK. By using a combination of mitogenic and enzymatic assays, we found that phosphorylation of the adjacent residue, serine 338, and, to a lesser extent, serine 339 is essential for the biological and enzymatic activities of Raf-1. Replacement of S338 with alanine blocked the ability of prenylated Raf-CX to transform Rat-1 fibroblasts. Similarly, the loss of S338-S339 in Raf-1 prevented protein kinase activation in COS-7 cells by either oncogenic Ras[V12] or v-Src. Consistent with phosphorylation of S338-S339, acidic amino acid substitutions of these residues partially restored transforming activity to Raf-CX, as well as kinase activation of Raf-1 by Ras[V12] or v-Src. Two-dimensional phosphopeptide mapping of wild-type Raf-CX and Raf-CX[A338A339] confirmed the presence of a phosphoserine-containing peptide with the predicted mobility in the wild-type protein which was absent from the mutant. This peptide could be quantitatively precipitated by an antipeptide antibody specific for the 18-residue tryptic peptide containing S338-S339 and was demonstrated to contain only phosphoserine. Phosphorylation of this peptide in Raf-1 was significantly increased by coexpression with Ras[V12]. These data demonstrate that Raf-1 residues 338 to 341 constitute a unique phosphoregulatory site in which the phosphorylation of serine and tyrosine residues contributes to the regulation of Raf by Ras, Src, and Ras-independent membrane localization.
Mol Cell Biol 1997 Aug
PMID:Phosphorylation of Raf-1 serine 338-serine 339 is an essential regulatory event for Ras-dependent activation and biological signaling. 923 8

Thrombopoietin (TPO) is the major regulator of both growth and differentiation of megakaryocytes. We previously showed that both functions can be generated by TPO in the megakaryoblastic cell line UT7, in which murine Mpl was introduced, and are independently controlled by distinct regions of the cytoplasmic domain of Mpl. Particularly, residues 71 to 94 of this domain (deleted in the mutant mpl delta3) were found to be required for megakaryocytic maturation but dispensable for proliferation. We show here that TPO-induced differentiation in UT7 cells is tightly dependent on a strong, long-lasting activation of the mitogen-activated protein kinase (MAPK) pathway. Indeed, (i) in UT7-mpl cells, TPO induced a strong activation of extracellular signal-regulated kinases (ERK) which was persistent until at least 4 days in TPO-containing medium; (ii) a specific MAPK kinase (MEK) inhibitor inhibited TPO-induced megakaryocytic gene expression; (iii) the Mpl mutant mpl delta3, which displayed no maturation activity, transduced only a weak and transient ERK activation in UT7 cells; and (iv) TPO-induced megakaryocytic differentiation in UT7-mpl delta3 cells was partially restored by expression of a constitutively activated mutant of MEK. The capacity of TPO to trigger a strong and prolonged MAPK signal depended on the cell in which Mpl was introduced. In BAF3-mpl cells, TPO triggered a weak and transient ERK activation, similar to that induced in UT7-mpl delta3 cells. In these cells, no difference in MAPK activation was found between normal Mpl and mpl delta3. Thus, depending on the cellular context, several distinct regions of the cytoplasmic domain of Mpl and signaling pathways may contribute to generate quantitative variations in MAPK activation.
Mol Cell Biol 1997 Sep
PMID:Control of thrombopoietin-induced megakaryocytic differentiation by the mitogen-activated protein kinase pathway. 927 77

Stress-activated protein kinases (SAPK; also known as JNK for c-Jun N-terminal kinase) phosphorylate Ser63 and Ser73 in the amino-terminus of the c-Jun protein and potentiate its transcriptional activity. We have analysed phosphorylation of GST fusion proteins containing the c-Jun N-terminal domain by lysates of Daudi human B lymphoblastoid cells stimulated with medium or anti-IgM. Crosslinking membrane IgM (mIgM) results in an increase in phosphorylation of GST-c-Jun (5-89) in an antibody dose-dependent manner. The kinase activity specifically phosphorylates the c-Jun N-terminal domain since it does not phosphorylate GST or GST-JunB. The activity preferentially phosphorylates the substrate that contains the sites for in vivo phosphorylation by SAPK/JNK and requires the delta domain of c-Jun, which is also required for SAPK/JNK activity. However, the c-Jun N-terminal kinase activity induced by mIgM ligation is not precipitatable with anti-SAPK/JNK antibodies. In addition, unlike SAPK/JNKs, the mIgM-dependent c-Jun N-terminal kinase activity is not detectable in assays for renaturable kinase activity (in-gel assay) or in assays that test activities that bind to c-Jun (solid-phase assay). The increased phosphorylation of c-Jun N-terminal domain in response to mIgM ligation is unlikely to be due to mIgM-activated ERKs as it was not suppressed by a selective MEK inhibitor. Thus, the mIgM-induced activity is distinct from the known SAPK/JNKs and may represent a novel mechanism for c-Jun phosphorylation in response to mIgM engagement in human B cells.
Mol Immunol 1997 Apr
PMID:Ligation of membrane IgM stimulates a novel c-Jun amino-terminal domain kinase activity in Daudi human B cells. 929 74

In 3T3-L1 fibroblasts, Ras proteins mediate both insulin-induced differentiation to adipocytes and its activation of cytosolic serine/threonine kinases, including Raf-1 kinase, mitogen-activated protein kinase (MAPK), and Rsk. Here, we report that insulin- and Ras-induced activation of MAPK is not required for the differentiation process and in fact antagonizes it. The treatment of 3T3-L1 preadipocytes with MEK-specific inhibitor PD98059 blocked insulin- and Ras-induced MAPK activation but had no effect on or slightly enhanced adipocytic differentiation. Tumor necrosis factor alpha (TNF-alpha), an inhibitor of insulin-stimulated adipogenesis, activated MAPK in 3T3-L1 cells. PD98059 treatment blocked MAPK activation by TNF-alpha and reversed the blockade of adipogenesis mediated by low (1 ng/ml) TNF-alpha concentrations. 3T3-L1 transfectants containing hyperactivated MEK1 or overexpressed MAPK displayed impaired adipocytic differentiation. PD98059 treatment also reversed the blockade of differentiation in MEK1 transfectants. These results indicate that MAPK does not promote but can contribute to inhibition of the process of adipocytic differentiation of 3T3-L1 cells.
Mol Cell Biol 1997 Oct
PMID:Mitogen-activated protein kinase activation is not necessary for, but antagonizes, 3T3-L1 adipocytic differentiation. 931 66

Mitogen-activated protein (MAP)/ERK kinase (MEK)1 and MEK2 are the upstream activators of the MAP kinases, ERK1 and ERK2. MEK1 and MEK2 are approximately 85% identical in sequence but have unique inserts in their C-terminal domains. MEK isoform-specific antibodies were used to examine expression and regulation of each enzyme. MEK1 and MEK2 were expressed in approximately equal amounts in several cell lines; in some, MEK1 was present in slight excess. Activation of tyrosine kinase-containing receptors, heterotrimeric G proteins, and protein kinase C enhanced the activities of both MEK isoforms in 293 and PC12 cells. AIF4-stimulated both MEK1 and MEK2 in PC12 cells expressing a dominant interfering Ras mutant that prevents nerve growth factor-dependent activation of the cascade. Carbachol also stimulated the pathway in these cells. Thus, in addition to their ability to activate Ras/Raf and the downstream ERK pathway, heterotrimeric G proteins also appear to trigger a Ras-independent mechanism to regulate this kinase cascade. In U373, Chinese hamster ovary (CHO), and INS-1 cells, MEK1 was activated by regulators of ERKs, while MEK2 was not. These data suggest that, like the MAP kinases ERK1 and ERK2, in some cell settings the two similar MEK isoforms are differentially regulated.
Mol Endocrinol 1997 Oct
PMID:Differential regulation of mitogen-activated protein/ERK kinase (MEK)1 and MEK2 and activation by a Ras-independent mechanism. 932 44

The HBx protein of hepatitis B virus (HBV) is a small transcriptional transactivator that is essential for infection by the mammalian hepadnaviruses and is thought to be a cofactor in HBV-mediated liver cancer. HBx stimulates signal transduction pathways by acting in the cytoplasm, which accounts for many but not all of its transcriptional activities. Studies have shown that HBx protein activates Ras and downstream Ras signaling pathways including Raf, mitogen-activated protein (MAP) kinase kinase kinase (MEK), and MAP kinases. In this study, we investigated the mechanism of activation of Ras by HBx because it has been found to be central to the ability of HBx protein to stimulate transcription and to release growth arrest in quiescent cells. In contrast to the transient but strong stimulation of Ras typical of autocrine factors, activation of Ras by HBx protein was found to be constitutive but moderate. HBx induced the association of Ras upstream activating proteins Shc, Grb2, and Sos and stimulated GTP loading onto Ras, but without directly participating in complex formation. Instead, HBx is shown to stimulate Ras-activating proteins by functioning as an intracellular cytoplasmic activator of the Src family of tyrosine kinases, which can signal to Ras. HBx protein stimulated c-Src and Fyn kinases for a prolonged time. Activation of Src is shown to be indispensable for a number of HBx activities, including activation of Ras and the Ras-Raf-MAP kinase pathway and stimulation of transcription mediated by transcription factor AP-1. Importantly, HBx protein expressed in cultured cells during HBV replication is shown to activate the Ras signaling pathway. Mechanisms by which HBx protein might activate Src kinases are discussed.
Mol Cell Biol 1997 Nov
PMID:Activation of Src family kinases by hepatitis B virus HBx protein and coupled signaling to Ras. 934 5


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