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 469 records · Page 26Linked to original sources

PubMed1986

Activated K-ras and N-ras oncogenes in primary renal mesenchymal tumors induced in F344 rats by methyl(methoxymethyl)nitrosamine.

Administration of methyl(methoxymethyl)nitrosamine to newborn Fischer 344 rats results in the preferential induction of renal tumors arising from the mesenchymal component of the kidney. DNA from a significant proportion of these tumors was capable of transforming NIH/3T3 cells. This report describes the renal tumor model, the detection of two different ras transforming genes in the kidney tumors (the N-ras oncogene in 1 and K-ras oncogene in 10 kidney tumors) and the characterization of DNA sequences specifying the transformed phenotype.

Animals↗

PubMedDate not supplied

[Initiation of the mouse sarcoma virus oncogene v-K-ras during the chemical treatment of proto-oncogene C-ras].

On the basis of the results obtained in the laboratories of Weinberg, Barbacid et al. describing the responsibility of one or another mutation in one of coding triplets of the ras family gene both for chemical and viral cancerogenesis, we draw the conclusion that it is impossible to distinguish these two causes of cancer appearance. It is shown that the cause of mice viral sarcoma v-K ras development from the cellular protooncogene C-ras may consist in the chemical reaction of one of coding triplet nucleotides with cancerogenic substance.

Animals↗

PubMed2005

Use of the Ras binding domain of c-Raf for biochemical and live-cell analysis of Ras activation.

Small modular GBDs (GTPase-binding domains) derived from GTPase-effector proteins are useful tools for the selective detection of the active GTP-loaded GTPase conformation, be it in biochemical assays or for imaging purposes. Use of GBD probes requires careful consideration of all features of the GDB-GTPase interaction. It is innate to the strong and specific interaction with the GTP-loaded GTPase, that GBDs will protect their partner GTPases from GAP (GTPase-activating protein) action. This feature is likely to cause an increase in cellular Ras-GTP levels, in particular in leucocytes and other cells with high steady-state Ras-GDP/GTP cycling rates. By the same token, high levels of GBD expression will interrupt GTPase-initiated signalling, with implications for the activation of the very same GTPase since feedback regulatory mechanisms can impinge on this process.

Animals↗

PubMed2003

Alternative splicing of the human proto-oncogene c-H-ras renders a new Ras family protein that trafficks to cytoplasm and nucleus.

We characterized a novel protein of the Ras family, p19 (H-RasIDX). The c-H-ras proto-oncogene undergoes alternative splicing of the exon termed IDX. We show that the alternative p19 mRNA is stable and as abundant as p21 (p21 H-Ras4A) mRNA in all of the human tissues and cell lines tested. IDX is spliced into stable mRNA in different mammalian species, which present a high degree of nucleotide conservation. Both the endogenous and the transiently expressed p19 protein are detected in COS-1 and HeLa cells and show nuclear diffuse and speckled patterns as well as cytoplasmic localization. In yeast two-hybrid assays, p19 did not interact with two known p21 effectors, Raf1 and Rin1, but was shown to interact with RACK1, a scaffolding protein that promotes multiprotein complexes in different signaling pathways. This observation suggests that p19 and p21 play differential and complementary roles in the cell.

3T3 Cells↗

PubMed1990

A common cellular pathway for v-mos and v-Ki-ras is not required for v-Ki-ras-induced tumorigenicity in a nonmalignant, v-mos-expressing revertant cell.

A revertant cell line was selected from Moloney sarcoma virus-transformed BALB/c cells after long-term treatment with type I interferon. Despite an actively transcribed and transfectable v-mos gene, these revertant cells were nontumorigenic in nude mice. The functionality of the mos protein was investigated, focusing on the alpha 2(1) collagen promoter regulation, which is known to be affected by mos-induced trans-acting factors. Both in transient expression assays and after stable integration into the cellular genome, the transfected alpha 2(1) collagen promoter fused to the cat reporter gene was activated in the revertant while being downregulated in the original transformed cells. In retransformation assays of the revertant by Moloney sarcoma virus strains homologous to the original transforming virus, no detectable change was noted in the in vitro phenotype or in tumorigenicity. These results reveal that the mos-directed factors were no longer effective on their specific targets. Thus, the R.MSVIF cell could be either an oncoprotein-deficient or a target-related revertant. Attempts at retransformation with unrelated sarcoma viruses bearing v-sis, v-fms, or v-fos oncogenes were also negative. In contrast, tumorigenicity was obtained with the unrelated Kirsten sarcoma virus without any change in the revertant morphology or collagen expression. These findings showed that the common pathway blocked by the reversion and shared by v-mos and v-ras was not required for ras-induced tumorigenicity.

Animals↗

PubMed2000

Human ras-converting enzyme (hRCE1) endoproteolytic activity on K-ras-derived peptides.

A human gene responsible for one of the steps in Ras post-translational modification and membrane localization, hRCE1, encodes a 35-kDa membrane-associated endoprotease. We examined hRCE1 activity using farnesylated 9 aa peptides with the core sequence, KSKTKC(farnesyl)VIM [(farnesyl) = (f)], from the C-terminus of K-Ras. We first demonstrated hRCE1 specificity in cleavage location and endoproteolysis. We then describe a direct fluorescent microtiter plate assay. We demonstrated that hRCE1 protease cleaved KSKTKC(f)VIM peptides between the C(f) and V positions, generating KSKTKC(f) and the corresponding tripeptides as products. We found that the sequence KSKTKC(f)VI was a better substrate for hRCE1 than KSKTKC(f)VIM. We also found that hRCE1 cleaved modified versions of KSKTKC(f)VIM that incorporated either MCA or ABZ fluorescent chromophores at the N-terminus, and quenching-group-containing amino acids at the V or M, but not the I, amino acid positions of VIM. The quenching-group-containing amino acids used were either Q(S) (dinitrophenyldiaminopropionic acid) or Q(L) (lysine epsilon-dinitrophenyl). Cleavage of KSKTKC(f)VIM and modified versions of this peptide by hRCE1 was initially evaluated by HPLC product resolution and quantitation. The hRCE1 cleavage of quenched peptides enabled us to directly monitor proteolytic activity in a 96-well microtiter fluorescent plate assay. The microtiter format assay was validated by its sensitivity to RPI, an inhibitor of prenyl protein protease. A direct fluorescent assay provides an effective tool for further characterization of this enzyme and also for detection of novel inhibitors.

Amino Acid Sequence↗

PubMed1994

Correlation between S-adenosyl-L-methionine content and production of c-myc, c-Ha-ras, and c-Ki-ras mRNA transcripts in the early stages of rat liver carcinogenesis.

gamma-Glutamyltranspeptidase (GGT)-positive and glutathione S-transferase (placental-GST-P) positive foci were induced in male Wistar rats by initiation with diethylnitrosamine (DENA), followed by selection and phenobarbital (PB). GGT- and GST-P-positive foci occupied 20-46% and 27-68% of liver parenchyma, respectively, 5-9 weeks after initiation. A high DNA synthesis was found in GGT-positive foci. Decrease in S-adenosyl-L-methionine (SAM) level and SAM/S-adenosylhomocysteine (SAH) ratio, and overall DNA hypomethylation occurred in the liver during the development of enzyme altered foci (EAF). These parameters underwent very small and transient changes in the liver of uninitiated rats at the 5th week, when EAF occupied 0.7-1.4% of the liver. At the 9th week, high RNA transcripts of c-myc, c-Ha-ras, and c-Ki-ras were found in the liver of initiated rats, but not in that of uninitiated rats. Immunohistochemical evaluation of c-myc gene product showed overexpression in GST-P-positive cells. SAM treatment of initiated rats caused inhibition of EAF growth, recovery of SAM/SAH ratio and DNA methylation, and decrease in protooncogene expression proportional to the dose and length of treatment. Liver SAM/SAH ratio was positively correlated with DNA methylation, and negatively correlated with transcript levels of the three protooncogenes. Thus, decrease in SAM/SAH ratio and DNA hypomethylation are early features of hepatocarcinogenesis promotion in rats fed a diet containing adequate lipotrope amounts, paralleled by overexpression of growth-related genes and rapid growth. Re-establishment of a physiologic SAM level makes it possible to inhibit protooncogene expression and EAF growth and to prevent late liver lesion development.

2-Acetylaminofluorene↗

PubMed2000

Treatment of v-Ki-ras-transformed SVC1 cells with low retinoic acid induces malignancy reversion associated with ras p21 down-regulation.

The effect of nontoxic, low concentrations (10(-8) M) of retinoic acid (RA) for a relatively long time (28 days) on a Kirsten ras-virus transformed cell line (Ki-SVC1), derived from the rat seminal vesicle epithelium, was investigated. In these experimental conditions, the cell treatment with RA induced a decrease of the proliferation rate, apoptosis and a marked reduction of both anchorage-independent growth and tumorigenicity. These biological responses were either preceded or associated with important changes in adenylate cyclase/protein kinase C signaling pathways, the activation of important apoptosis-linked genes and a marked decrease of the v-Ki-ras p21 protein. The significance of these findings is discussed.

Adenylyl Cyclases↗

PubMed2001

Ras effectors: buying shares in Ras plc.

The debate over whether activated Ras can regulate phosphoinositide-specific phospholipase C (PLC) has been contentious and at times heated. The argument may be resolved by the recent identification of a novel Ras-regulated PLC, but some unexpected properties of this protein are sure to stimulate further controversy.

Humans↗

PubMed1999

The pre-hydrolysis state of p21(ras) in complex with GTP: new insights into the role of water molecules in the GTP hydrolysis reaction of ras-like proteins.

BACKGROUND: In numerous biological events the hydrolysis of guanine triphosphate (GTP) is a trigger to switch from the active to the inactive protein form. In spite of the availability of several high-resolution crystal structures, the details of the mechanism of nucleotide hydrolysis by GTPases are still unclear. This is partly because the structures of the proteins in their active states had to be determined in the presence of non-hydrolyzable GTP analogues (e.g. GppNHp). Knowledge of the structure of the true Michaelis complex might provide additional insights into the intrinsic protein hydrolysis mechanism of GTP and related nucleotides. RESULTS: The structure of the complex formed between p21(ras) and GTP has been determined by X-ray diffraction at 1.6 A using a combination of photolysis of an inactive GTP precursor (caged GTP) and rapid freezing (100K). The structure of this complex differs from that of p21(ras)-GppNHp (determined at 277K) with respect to the degree of order and conformation of the catalytic loop (loop 4 of the switch II region) and the positioning of water molecules around the gamma-phosphate group. The changes in the arrangement of water molecules were induced by the cryo-temperature technique. CONCLUSIONS: The results shed light on the function of Gln61 in the intrinsic GTP hydrolysis reaction. Furthermore, the possibility of a proton shuffling mechanism between two attacking water molecules and an oxygen of the gamma-phosphate group can be proposed for the basal GTPase mechanism, but arguments are presented that render this protonation mechanism unlikely for the GTPase activating protein (GAP)-activated GTPase.

Crystallography, X-Ray↗

PubMed1999

Molecular dynamics analysis of the structures of ras-guanine nucleotide exchange protein (SOS) bound to wild-type and oncogenic ras-p21. Identification of effector domains of SOS.

The X-ray crystal structure of the ras oncogene-encoded p21 protein bound to SOS, the guanine nucleotide exchange-promoting protein, has been determined. We have undertaken to determine if there are differences between the three-dimensional structures of SOS bound to normal and oncogenic (Val 12-p21) proteins. Using molecular dynamics, we have computed the average structures for both complexes and superimposed them. We find four domains of SOS that differ markedly in structure: 631-641, 676-691, 718-729, and 994-1004. Peptides corresponding to these sequences have been synthesized and found to be powerful modulators of oncogenic p21 in cells as described in an accompanying paper. We find that the SOS segment from 809-815 makes contacts with multiple domains of ras-p21 and can facilitate correlated conformational changes in these domains.

Models, Molecular↗

PubMed1997

Molecular dynamics on complexes of ras-p21 and its inhibitor protein, rap-1A, bound to the ras-binding domain of the raf-p74 protein: identification of effector domains in the raf protein.

We have computed the average structures for the ras-p21 protein and its strongly homologous inhibitor protein, rap-1A, bound to the ras-binding domain (RBD) of the raf protein, using molecular dynamics. Our purpose is to determine the differences in structure between these complexes that would result in no mitogenic activity of rap-1A-RBD but full activity of p21-RBD. We find that despite the similarities of the starting structures for both complexes, the average structures differ considerably, indicating that these two proteins do not interact in the same way with this vital target protein. p21 does not undergo major changes in conformation when bound to the RBD, while rap-1A undergoes significant changes in structure on binding to the RBD, especially in the critical region around residue 61. The p21 and rap-1A make substantially different contacts with the RBD. For example, the loop region from residues 55-71 of rap-1a makes extensive hydrogen-bond contacts with the RBD, while the same residues of p21 do not. Comparison of the structures of the RBD in both complexes reveals that it undergoes considerable changes in structure when its structure bond to p21 is compared with that bound to rap-1A. These changes in structure are due to displacements of regular structure (e.g., alpha-helices and beta-sheets) rather than to changes in the specific conformations of the segments themselves. Three regions of the RBD have been found to differ significantly from one another in the two complexes: the binding interface between the two proteins at residues 60 and 70, the region around residues 105-106, and 118-120. These regions may constitute effector domains of the RBD whose conformations determine whether or not mitogenic signal transduction will occur.

GTP-Binding Proteins↗

PubMed1989

Instability of expression of major histocompatibility antigens in fibroblasts expressing activated ras oncogene: constitutive and interferon-gamma induced class I and class II antigens in a series of clonal isolates of murine fibroblasts transformed by v-Ki-ras.

We have examined the expression of major histocompatibility complex (MHC) antigens, constitutive or induced with interferon gamma (IFN-gamma), in a line of C3H mouse embryo fibroblasts (C3H 201) transformed with a helper-virus-free preparation of the Kirsten strain of murine sarcoma virus. C3H 201 cells expressed some class I antigen (H-2Kk) in the absence of added interferon, unlike the parental C3H 10T1/2 cells from which they were derived. However, this declined with (in vitro) passage level after transformation. Treatment with IFN-gamma induced very high expression of H-2Kk at all passage levels. There was no constitutive expression of class II antigen (I-Ak); however, this could be induced by IFN-gamma. Inducibility of I-Ak was found also to be related to the number of passages after transformation; at early passage levels after transformation more I-Ak was induced than after the cells had been allowed to grow for several passages, until at high passage levels little or no I-Ak was induced. This was not due to the presence of a subpopulation of untransformed cells since when the cells were cloned shortly after infection all the resulting clones were transformed. In addition, IFN-gamma at any passage level induced clearly less I-Ak than was found in C3H 10T1/2 cells, in which I-Ak inducibility was high and stable. Twenty-one clones were derived from C3H 201 cells at early passage (less than 8) either from soft agar or from liquid culture. These clones showed a wide variation in MHC antigen phenotype. Many expressed H-2Kk in the absence of IFN-gamma, and all were strongly inducible for H-2Kk. None showed I-Ak in the absence of IFN-gamma. All but two expressed I-Ak after IFN-gamma treatment but, with four exceptions, clearly less than the untransformed line. Four clones derived at late passage (40) resembled the late passage line. The expression of the ras oncogene and tumorigenicity was studied in representative clones; there was no obvious correlation with MHC phenotype, nor with the method of cloning. We conclude from these studies that the expression of MHC antigens by fibroblasts expressing the v-Ki-ras oncogene, either with or without exposure to interferon gamma, is unstable, varying with the number of cell generations from transformation and from clone to clone.

Animals↗

PubMed2004

Ras-dependent induction of HIF-1alpha785 via the Raf/MEK/ERK pathway: a novel mechanism of Ras-mediated tumor promotion.

Hypoxia-inducible factor-1alpha (HIF-1alpha) plays crucial roles in tumor promotion by transactivating approximately 60 kinds of its target genes. Recently, we reported a novel splice variant HIF-1alpha(785), which is regulated primarily by phorbol ester. This variant can be stabilized under normoxic conditions because it loses an acetylation site Lys532. Its expression was found to promote xenografted tumor growth in nude mice. We here found that the Ras oncogene regulates HIF-1alpha(785) expression via the Raf/MEK/ERK pathway, and that both phorbol ester and epidermal growth factor also induced HIF-1alpha(785) via the same pathway. We also identified the nonhypoxic regulatory domain responsible for phorbol ester-induced HIF-1alpha(785) expression. These results imply that HIF-1alpha(785) may play an important role in tumor promotion mediated by the Ras oncogene, phorbol ester or tumor growth factors.

Animals↗

PubMed1999

In budding yeast, reactive oxygen species induce both RAS-dependent and RAS-independent cell cycle-specific arrest.

The role of mild oxidative stresses elicited by diethylmaleate (DEM)-induced glutathione depletion in the progression of the yeast cell cycle has been investigated. We found that different wild-type strains are sensitive to oxidative stresses induced by similar DEM doses: approximately 1 mM on YPD plates, 5-10 mM in shaken flasks. At lower doses, DEM caused a transient decrease in growth rate, largely because of a decreased G1-to-S transition. Treatment with higher DEM doses leads to complete growth arrest, with most cells found in the unbudded G1 phase of the cell cycle. DEM treatment resulted in transcriptional induction of stress-responsive element (STRE)-controlled genes and was relieved by treatment with the antioxidant N-acetyl cysteine. Reciprocal shift experiments with cdc25 and cdc28 mutants showed that the major cell cycle arrest point was located in the Start area, at or near the CDC25-mediated step, before the step mediated by the CDC28 cyclin-dependent kinase. The DEM-induced G1 arrest requires a properly regulated RAS pathway and can be bypassed by overexpressing the G1-specific cyclin CLN2. However, cells with either a deregulated RAS pathway or overexpressing CLN2 failed to grow and arrested as budded cells, indicating that a second DEM-sensitive cell cycle step exists.

Acetylcysteine↗

PubMed2001

Mechanisms of inhibition of the Ras-MAP kinase signaling pathway in 30.7b Ras 12 cells by tea polyphenols (-)-epigallocatechin-3-gallate and theaflavin-3,3'-digallate.

Our previous study showed that tea polyphenols inhibited MAP kinase and AP-1 activities in mouse epidermal JB6 cells and the corresponding H-ras-transformed cell line 30.7b Ras 12. The present study investigated the mechanisms of this inhibition. The cells were incubated with (-)-epigallocatechin-3-gallate (EGCG) or theaflavin-3,3'-digallate (TFdiG) (20 mM) for different times, and the cell lysate was analyzed by immunoblotting. EGCG treatment decreased the levels of phospho-Erk1/2 and -MEK1/2 time-dependently (by 60% at 60 min). TFdiG lowered their levels by 38%-50% at 15 min. TFdiG effectively decreased total Raf-1 protein levels, most likely through lysosomal degradation. EGCG did not affect protein levels or the activity of Raf-1 significantly but decreased its association with MEK1 as determined by co-immunoprecipitation. In addition, EGCG and TFdiG (10 mM) inhibited the phosphorylation of Elk-1 by isolated phospho-Erk1/2 in vitro. This inhibition of Erk1/2 activity is Elk-1 concentration-dependent and ATP concentration-independent, which suggests that EGCG and TFdiG interfere with the binding of the protein substrate to the kinase. The presently demonstrated specific mechanisms of inhibition of MAP kinases by EGCG and TFdiG may help us to understand the effects of tea consumption on cancer, inflammatory diseases, and cardiovascular diseases.

Adenosine Triphosphate↗

PubMed1999

Characterization of the hinges of the effector loop in the reaction pathway of the activation of ras-proteins. Kinetics of binding of beryllium trifluoride to V29G and I36G mutants of Ha-ras-p21.

This work experimentally confirms the pathway of activation of Ha-ras-p21, which was calculated by the method of Targeted Molecular Dynamics (TMD) (Díaz JF, Wroblowski B, Schlitter J, Engelborghs Y, 1997a, Proteins Struct Funct Genet 28:434-451). The process can be studied experimentally by analyzing the binding of BeF3- to the GDP complex of the active fluorescent mutant Y32W (Díaz JF, Sillen A, Engelborghs Y, 1997b, J Biol Chem 227:23138-23143). Two mutants, V29G and 136G, have been constructed at both sides of the effector loop of the active fluorescent mutant. This was done to check the proposed reaction pathway and to provide further insight into the mechanism of the activation of ras proteins. Both mutations accelerate the conformational isomerization with two orders of magnitude, demonstrating convincingly the role of these residues as hinges of the effector loop in one or more of the transitions of the conformational change. These results provide experimental support to the pathway calculated by TMD analysis.

Beryllium↗

PubMed1990

Mutational activation of H-ras and K-ras genes is absent in N-nitroso-N-methylurea-induced liver tumors in rats.

We examined mutational activation of H- and K-ras genes in hyperplastic nodules and hepatocellular carcinomas induced by N-nitroso-N-methylurea or diethylnitrosamine using the polymerase chain reaction, followed by dot-blot hybridization. No mutational activation was detected in these hepatic lesions. The results indicate that low incidence of the activation of H- and K-ras genes in rat liver tumors is due to the organ specificity rather than the nature of the carcinogens used.

Animals↗