SEARCH · PubMed Health

Results for “RAS”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 397 records · Page 22Linked to original sources

PubMed1994

Hepatitis B virus HBx protein activates Ras-GTP complex formation and establishes a Ras, Raf, MAP kinase signaling cascade.

Hepatitis B virus produces a small (154-amino acid) transcriptional transactivating protein, HBx, which is required for viral infection and has been implicated in virus-mediated liver oncogenesis. However, the molecular mechanism for HBx activity and its possible influence on cell proliferation have remained obscure. A number of studies suggest that HBx may stimulate transcription by indirectly activating transcription factors, possibly by influencing cell signaling pathways. We now present biochemical evidence that HBx activates Ras and rapidly induces a cytoplasmic signaling cascade linking Ras, Raf, and mitogen-activated protein kinase (MAP kinase), leading to transcriptional transactivation. HBx strongly elevates levels of GTP-bound Ras, activated and phosphorylated Raf, and tyrosine-phosphorylated and activated MAP kinase. Transactivation of transcription factor AP-1 by HBx is blocked by inhibition of Ras or Raf activities but not by inhibition of Ca(2+)- and diacylglycerol-dependent protein kinase C. HBx was also found to stimulate DNA synthesis in serum-starved cells. The hepatitis B virus HBx protein therefore stimulates Ras-GTP complex formation and promotes downstream signaling through Raf and MAP kinases, and may influence cell proliferation.

Animals↗

PubMed2002

Ras and TGF[beta] cooperatively regulate epithelial cell plasticity and metastasis: dissection of Ras signaling pathways.

Multistep carcinogenesis involves more than six discrete events also important in normal development and cell behavior. Of these, local invasion and metastasis cause most cancer deaths but are the least well understood molecularly. We employed a combined in vitro/in vivo carcinogenesis model, that is, polarized Ha-Ras-transformed mammary epithelial cells (EpRas), to dissect the role of Ras downstream signaling pathways in epithelial cell plasticity, tumorigenesis, and metastasis. Ha-Ras cooperates with transforming growth factor beta (TGFbeta) to cause epithelial mesenchymal transition (EMT) characterized by spindle-like cell morphology, loss of epithelial markers, and induction of mesenchymal markers. EMT requires continuous TGFbeta receptor (TGFbeta-R) and oncogenic Ras signaling and is stabilized by autocrine TGFbeta production. In contrast, fibroblast growth factors, hepatocyte growth factor/scatter factor, or TGFbeta alone induce scattering, a spindle-like cell phenotype fully reversible after factor withdrawal, which does not involve sustained marker changes. Using specific inhibitors and effector-specific Ras mutants, we show that a hyperactive Raf/mitogen-activated protein kinase (MAPK) is required for EMT, whereas activation of phosphatidylinositol 3-kinase (PI3K) causes scattering and protects from TGFbeta-induced apoptosis. Hyperactivation of the PI3K pathway or the Raf/MAPK pathway are sufficient for tumorigenesis, whereas EMT in vivo and metastasis required a hyperactive Raf/MAPK pathway. Thus, EMT seems to be a close in vitro correlate of metastasis, both requiring synergism between TGFbeta-R and Raf/MAPK signaling.

Animals↗

PubMed1992

The tumour promoters dieldrin and phenobarbital increase the frequency of c-Ha-ras wild-type, but not of c-Ha-ras mutated focal liver lesions in male C3H/He mice.

The frequency and pattern of mutations at codon 61 of the c-Ha-ras protooncogene were analysed in glucose-6-phosphatase-deficient hepatic lesions of male C3H/He mice occurring either spontaneously or after continuous treatment with 10 p.p.m. dieldrin or 500 p.p.m. phenobarbital (PB) in their diet. At 52 weeks after start of promoter administration, enzyme-altered liver lesions had developed in 41% (15/37) of untreated control mice and in 67% (10/15) and 63% (10/16) of mice treated with dieldrin or PB respectively. The average numbers of focal lesions per mouse were 0.57 in the control, 1.5 in the dieldrin and 1.0 in the PB group. Lesions were punched out from frozen liver sections and used for mutation analysis by allele-specific oligonucleotide hybridization following in vitro amplification of DNA via polymerase chain reaction. In the control group, 12 out of 21 liver lesions (57%) showed c-Ha-ras mutations, while five out of 23 (22%) and four out of 16 (25%) lesions were mutated in the dieldrin and PB groups. Taking the different numbers of animals in the three experimental groups into account, our data indicate that the tumour promoters increased the frequency of c-Ha-ras wild-type but not of c-Ha-ras mutated focal liver lesions, suggesting that the mutations had occurred spontaneously and were not related to treatment. Since c-Ha-ras mutations were found to be frequent in large but infrequent in small hepatocellular lesions, these mutations may represent in livers of C3H/He mice an endogenous promoting principle that provides a selective growth advantage to the mutated progenitor cells.

Amino Acid Sequence↗

PubMed2003

Vav mediates Ras stimulation by direct activation of the GDP/GTP exchange factor Ras GRP1.

Here we describe a new signaling cross-talk between the Vav/Rac1 and Ras pathways that is established through the stimulation of RasGRP1, an exchange factor for Ras subfamily GTPases. This interaction is crucial for Ras activation in lymphoid cells, since this GTPase cannot become activated in the absence of Vav proteins. The activation of RasGRP1 requires both the generation of diacylglycerol via phospho lipase C-gamma and the induction of actin polymerization, two responses induced by Vav and Rac1 that facilitate the translocation of RasGRP1 to juxtamembrane areas of the cell. Consistent with this, the cross-talk can be activated by tyrosine-phosphorylated wild-type Vav, oncogenic Vav and constitutively active Rac1. Conversely, Ras activation can be blocked in lymphocytes and ectopic systems using inhibitors affecting either phospholipase C-gamma or F-actin polymerization. These results indicate that a relay mechanism exists in lymphoid and other cells helping in the generation of robust signaling responses by the Rac/Rho and Ras pathways upon receptor engagement.

Actins↗

PubMed2003

Overexpression of kinase suppressor of Ras upregulates the high-molecular-weight tropomyosin isoforms in ras-transformed NIH 3T3 fibroblasts.

The down-regulation of the high-molecular-weight isoforms of tropomyosin (TM) is considered to be an essential event in cellular transformation. In ras-transformed fibroblasts, the suppression of TM is dependent on the activity of the Raf-1 kinase; however, the requirement for other downstream effectors of Ras, such as MEK and ERK, is less clear. In this study, we have utilized the mitogen-activated protein kinase scaffolding protein Kinase Suppressor of Ras (KSR) to further investigate the regulation of TM and to clarify the importance of MEK/ERK signaling in this process. Here, we report that overexpression of wild-type KSR1 in ras-transformed fibroblasts restores TM expression and induces cell flattening and stress fiber formation. Moreover, we find that the transcriptional activity of a TM-alpha promoter is decreased in ras-transformed cells and that the restoration of TM by KSR1 coincides with increased transcription from this promoter. Although ERK activity was suppressed in cells overexpressing KSR1, ERK inhibition alone was insufficient to upregulate TM expression. The KSR1-mediated effects on stress fiber formation and TM transcription required the activity of the ROCK kinase, because these effects could be suppressed by the ROCK inhibitor, Y27632. Overexpression of KSR1 did not directly regulate ROCK activity, but did permit the recoupling of ROCK to the actin polymerization machinery. Finally, all of the KSR1-induced effects were mediated by the C-terminal domain of KSR1 and were dependent on the KSR-MEK interaction.

3T3 Cells↗

PubMed1994

Synergistic activation by Ras and 14-3-3 protein of a mitogen-activated protein kinase kinase kinase named Ras-dependent extracellular signal-regulated kinase kinase stimulator.

We have identified, in Xenopus oocyte cytosol, a protein kinase named REKS (Ras-dependent extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase kinase (MEK) stimulator), which phosphorylates and activates recombinant ERK2 through recombinant MEK in a recombinant GTP gamma S (guanosine 5'-(3-O-thio)triphosphate)-Ras-dependent manner. We show here that this REKS activity is synergistically enhanced by a combination of mammalian recombinant GTP gamma S-KiRas and 14-3-3 protein purified from rat brain. 14-3-3 protein is known to activate tyrosine and tryptophan hydroxylases, to modulate the protein kinase C activity, to stimulate secretion, and to show phospholipase A2 activity per se. 14-3-3 protein did not affect the MEK activity. 14-3-3 protein neither interacted with Ki-Ras nor affected the neurofibromin activity to stimulate the GTPase activity of Ki-Ras under the conditions where the recombinant N-terminal fragment of c-Raf-1 inhibited it. These results suggest that 14-3-3 protein has an additional function in the regulation of the Ras-MEK-ERK cascade pathway through the activation of REKS.

14-3-3 Proteins↗

PubMed1988

Analysis of metastatic competence of mouse bladder carcinoma cells after transfection with activated Ha-ras or N-ras oncogenes.

Transfection of the Ha-ras oncogene into a low metastatic epithelial cell line resulted in the acquirement of significantly increased metastatic capacity. This alteration in metastatic competence of a carcinoma line in a syngeneic system seemed to be a selective change and was not affected by parameters such as tumor latency period or local tumor growth. Transfection of the selection marker vectors with normal cellular DNA or with the N-ras gene did not lead to significantly increased metastatic capacity. Analysis of metastatic variants after oncogene transfection and in vivo selection showed integration of N-ras, but not of Ha-ras oncogenes. A possible role for the Ha-ras oncogene in the initial steps of metastasis will be discussed.

Animals↗

PubMed1986

Expression of the proto-oncogenes C-H-ras and N-ras in early second trimester human fetal tissues.

Tissues from human fetuses of 12-18 weeks gestational age were examined for expression of c-oncogenes. Dot blot hybridization showed that N-ras and C-H-ras were expressed to varying degrees in all tissues. Some of the tissues were further examined by Northern blot hybridization, and transcripts of approximately 2.2 Kb and 5.2 Kb were detected using N-ras cDNA probe. A further transcript of approximately 1.4 Kb was observed when the same blot was hybridized to C-H-ras cDNA probe. No hybridization was detected between the fetal tissues and c-K-ras, c-myc, v-erb, c-fos, c-sis and c-src.

Fetus↗

PubMed1988

c-H-ras and c-K-ras gene hypomethylation in the livers and hepatomas of rats fed methyl-deficient, amino acid-defined diets.

The extent of methylation of the c-H-ras and c-K-ras oncogenes was compared in neoplastic and preneoplastic livers of rats fed one of several methyl-deficient, amino acid-defined diets for 18 months, with or without a preceding initiating dose of diethylnitrosamine (DEN). The restriction endonucleases MspI, HpaII, HhaI, PaeR71 and XhoI were used for studying the extent and pattern of DNA methylation. The results indicated that both c-H-ras and c-K-ras oncogenes were hypomethylated in all DNA samples derived from both neoplastic and preneoplastic livers of rats fed any of the methyl-deficient diets used, regardless of whether or not the rats had received an initiating dose of DEN. It thus appears that dietary methyl deficiency does indeed lead to hypomethylation of ras genes in the DNAs of the resulting tumors. However, the significance of this hypomethylation in the tumorigenic process is not clearly understood.

Amino Acids↗

PubMed1988

Activated c-K-ras and c-N-ras oncogenes in 3-methylcholanthrene-induced BALB/c fibrosarcomas.

DNAs from fourteen fibrosarcomas induced by 3-methylcholanthrene (MCA) in BALB/c mice were analyzed for the presence of transforming oncogenes following transfection on NIH3T3 cells. Six transfection-positive DNAs contained an activated ras gene: four c-K-ras and two c-N-ras. These results demonstrate that c-K-ras is not the only oncogene of the ras family activated in MCA-induced murine fibrosarcomas as was previously indicated.

Animals↗

PubMed2004

Quantification of absolute Ras-GDP/GTP levels by HPLC separation of Ras-bound [(32)P]-labelled nucleotides.

Ras guanine nucleotide binding protein (GTPase) activation is a widely assessed readout in cell biological studies. We describe an improved approach for the quantitative analysis of total GDP and GTP bound to Ras. The present method involves HPLC separation and online detection/quantitation of Ras-bound [(32)P]-labelled GDP/GTP. As compared to standard approaches that are time consuming and/or provide only semi-quantitative data, this technique allows the rapid processing of large numbers of samples for the quantitative determination of Ras-bound GDP and GTP.

Animals↗

PubMed2001

Paired human fibrosarcoma cell lines that possess or lack endogenous mutant N-ras alleles as experimental model for Ras signaling pathways.

We present here a human cell model for examination of mutant N-ras function. The HT1080 human fibrosarcoma cell line is pseudodiploid and contains a single endogenous mutant N-ras allele. MCH603c8 cells are a variant of HT1080 cells, in which the mutant allele has been deleted. The two cell lines differ dramatically in the constitutive levels of activation of downstream members of the Ras signaling pathways, and in biological features of transformation and tumorigenicity. Downregulation or activation of individual Ras-dependent pathways can be accomplished via transfection of dominant negatives or activated mutant cDNAs into HT1080 and MCH603c8 cells, respectively. The biochemical and biological consequences of expression of these mutant cDNAs can be assessed. There are dramatic effects on both the transformed and tumorigenic phenotype, depending on the cell line and mutant cDNA that is transfected.

Actins↗

PubMed1997

Ras-GRF, the activator of Ras, is expressed preferentially in mature neurons of the central nervous system.

In rodents, the Ras-specific guanine-nucleotide exchange factor (Ras-GRF) is expressed in different areas of the brain and, at a reduced level, also in the spinal cord. No expression of the 140 kDa Ras-GRF was detected in dorsal root ganglia and all other tissues tested. Analysis of primary cultures derived from brain reveals that this exchange factor is only present in neurons of the central nervous system. In primary hippocampal cultures, the expression of Ras-GRF increases in parallel with the onset of a neuronal network and in the whole brain it increases sharply after birth.

Animals↗

PubMed1990

Neutron radiation can activate K-ras via a point mutation in codon 146 and induces a different spectrum of ras mutations than does gamma radiation.

Neutron radiation is known to produce tumors in animals and cause cell transformation. We have developed a protocol to efficiently induce thymic lymphomas in RF/J mice by a single acute dose of neutron irradiation. Activated ras genes were detected in 17% (4 of 24) of the tumors analyzed. One of the tumors contained a K-ras gene activated by a point mutation in codon 146. Activating ras mutations at position 146 have not been previously detected in any known human or animal tumors. The spectrum of ras mutations detected in neutron radiation-induced thymic lymphomas was different from that seen in thymic lymphomas induced by gamma radiation in the same strain of mice. These results may have important implications for the mechanisms by which different types of radiation damage DNA.

Animals↗

PubMedDate not supplied

Inhibition of the membrane localization of p21 ras proteins by lovastatin in tumor cells possessing a mutated N-ras gene.

Mutated ras genes are found in a variety of human tumors. For biological activity the gene product p21 ras needs to be bound to the cell membrane by a farnesyl residue. Treatment of tumor cells with lovastatin reduces the availability of farnesyl pyrophosphate for the modification of the ras proteins. The membrane localization of p21 ras has been reduced by 30-36% after the tumor cells have grown in the presence of 10 microM lovastatin for 7 days. The extent of the inhibition depends on the growth kinetics of the cell lines.

Acute Disease↗

PubMed2004

Destruxin E, a cyclodepsipeptide antibiotic, reduces cyclin D1 levels and inhibits anchorage-independent growth of v-Ki-ras-expressed pMAM-ras-REF cells.

Destruxin E (DE), a cyclodepsipeptide isolated from fermentation broths of Metarhizium sp. MA324, inhibited the growth of v-Ki-ras-expressed pMAM-ras-REF (rasREF) cells in the suspension (anchorage-independent) culture (a) more strongly than that in the substratum-attached (anchorage-dependent) culture (b) or that of v-Ki-ras-unexpressed pMAM-ras-REF (REF) cells in the substratum-attached culture (c); the IC(50) values of DE were 0.07 microM (a), 0.4 microM (b), and 1.2 microM (c). DE arrested G1 phase cell cycle progression of rasREF cells in the substratum-attached culture (b). In rasREF cells treated with DE for 72 h in suspension culture (a), the levels of cyclin D1, cyclin A, p27(Kip1), and hyperphosphorylated Rb were decreased, but the levels of cdk4, cdk6, cdk2, p16(INK4a), and p21(Cip1) were not affected. Among these effects, the decrease in cyclin D1 was prominent. DE decreased the level of cyclin D1 in rasREF cells in the suspension culture (a) at 0.1 microM and in the substratum-attached culture (b) at 1 microM, while the level of cyclin D1 in REF cells in the substratum-attached culture (c) was not decreased at 1 microM. The extent of growth inhibition correlated with the decrease in cyclin D1. The level of cyclin D1 mRNA of rasREF cells in the suspension culture (a) was also decreased by DE. DE decreased cyclin D1 mRNA, resulting in inhibition of anchorage-independent growth of rasREF cells.

Animals↗

PubMed1994

Cdc25p, the guanine nucleotide exchange factor for the Ras proteins of Saccharomyces cerevisiae, promotes exchange by stabilizing Ras in a nucleotide-free state.

In Saccharomyces cerevisiae, adenylate cyclase activity is controlled by Ras1p and Ras2p. Activation of the Ras proteins is in turn controlled by the GTPase-activating proteins (GAPs), Ira1p and Ira2p, and the guanine nucleotide exchange factor (GNEF), Cdc25p. We have characterized Cdc25p enzymologically in order to gain information about the mechanism of Cdc25p-mediated guanine nucleotide exchange and to appreciate how the activity of a GNEF is integrated as a part of a basic molecular switch module consisting of Ras, GNEF, and GAP. Using Ras2p and a catalytic fragment of Cdc25p, both expressed in and purified from Escherichia coli, we determined that Cdc25p has a Km for Ras2p-GDP of 160 nM and a maximal rate of 0.20 s-1. The Km of Cdc25p for Ras2p complexed to GTP is 3-fold greater than that for Ras2p complexed to GDP. The Km of free GDP is about 2-fold higher than the Km of free GTP. This suggests that Cdc25p activates Ras2p primarily by equilibrating Ras2p with the pool of free guanine nucleotides in the cell rather than by driving Ras2p inexorably into the activated state. This renders Ras activation potentially subject to energy charge fluctuations in the cell. The free guanine nucleotide affects kcat, indicating that the rate-limiting step is nucleotide association. Finally, we demonstrated that dominant negative alleles of Ras2p are potent competitive inhibitors of Cdc25p. These data, in conjunction with the kinetic data, are consistent with the hypothesis that Cdc25p catalyzes guanine nucleotide exchange by stabilizing a nucleotide-free intermediate of Ras.

Cell Cycle Proteins↗

PubMed1993

[Mutational activation of H-ras and K-ras (Codon 12,61) genes in 7,12 dimethylbenz(a)anthracene induced rat ovarian tumors].

We had studied the histogenesis and p21 expression in ovarian tumor induced by 7,12-dimethylbenz(a)anthracene (DMBA). The present study was done to elucidate point mutation of the ras gene in tumors. Cases with immunohistological p21 expression; 3 adenomas, 5 adenocarcinomas and 2 sarcomas were examined by polymerase chain reaction (PCR). 1) The products of PCR were confirmed the presence of amplified sequences by electrophoresis. 2) No point mutation of H-ras (Codon 12,61) or K-ras (Codon 12,61) genes was found in the ovarian tumors induced by DMBA. 3) These results suggested that the mutational activation of ras genes in tumorigenesis was dependent on another mechanism.

9,10-Dimethyl-1,2-benzanthracene↗