Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The c-Ha-ras oncogene has been implicated as a causative agent in the development of tumors in humans as well as mice. The molecular nature of the ras-induced tumorigenic process remains unclear, however. To address this question directly we have constructed a cell line which carries a zinc-inducible metallothionein-ras hybrid oncogene, transformant 212. Upon exposure to zinc for 24-48 hr, 212 cells assume a highly transformed morphology, concomitant with the induction of ras-expression. Natural killer cells constitute a subpopulation of lymphoid effector cells which have for a long time been hypothesized to be involved in the earliest stages of antitumor surveillance. Central to this hypothesis is the prediction that NK sensitivity arises during cellular transformation. By carrying out cytotoxicity assays against the 212 transformant, we showed that, indeed, increased sensitivity to NK-mediated lysis correlated with expression of the ras oncogene, which is consistent with the above hypothesis. We then addressed the question of the biochemical mechanism of ras-induced transformation. Owing to their similarity to G proteins, regulatory elements interposed between cell-surface receptors and their effector enzymes, it has been postulated that p21, the ras oncogene protein, mediates its transforming effects by constitutive activation of proliferative signal transduction pathways. We studied the effect of ras expression on the regulation of adenylate cyclase (A.C.), key enzyme of one such major pathway. We found that ras expression correlated with a dampening of responsiveness of A.C. to several stimuli, including hormones such as isoproterenol and other agents such as GMP-PNP, forskolin and fluoride-ion. Accumulation of cAMP as measured by RIA in intact cells, as basal or in response to stimulation of A.C. activity with forskolin, was also decreased (approximately 10-fold) with ras expression. Because the regulation of calcium, another important second messenger is dependent, in part, upon cAMP and GTP-binding proteins, we investigated the possible influence of ras expression on the intracellular concentration of calcium. Steady-state intracellular free Ca2+ concentration, as measured by fluorimetry, was indeed increased by approximately 50-125% in association with ras expression. Finally, we studied the possible influence of p21ras on protein kinase C (PKC), which is a key enzyme in the important signal transduction pathway of phosphatidylinositol lipid turnover. We assessed PKC activity directly, in a cell-free system, by measuring the ability of the enzyme to transfer radiolabelled phosphate from gamma-32P-ATP to histone, and exogenous substrate.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:The cell biology of ras-induced transformation: insights from studies utilizing an inducible hybrid oncogene system. 284 54

Certain tumour cells contain activated ras genes that code for 21 000 dalton proteins (p21). These proteins associate with the inner face of the plasma membrane and bind guanine nucleotides specifically. In order to determine whether p21s have functions similar to other GTP binding proteins, we investigated the regulation, by guanine nucleotides, of adenylate cyclase (AC) activity in membrane preparations isolated from fibroblasts (C127) transformed by a temperature sensitive mutant of Kirsten sarcoma virus (Ts 371). The degree of AC stimulation by GMP P(NH)P increased when these cells were shifted from the permissive temperature (33 degrees C) to the non-permissive temperature (39 degrees C). This effect was more pronounced at low Mg++ and low GMP P(NH)P concentrations. AC stimulation remained unchanged in rat fibroblasts infected with a temperature sensitive mutant of Rous Sarcoma virus. AC activity was depressed in C127 cells infected with wild type KiMSV. Our data illustrate the feasibility of correlating alterations in the AC system with ras gene expression and using such experimental approaches to elucidate the physiological functions of the p21 proteins.
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PMID:Modulation of adenylate cyclase by guanine nucleotides and Kirsten sarcoma virus mediated transformation. 298 13

The Ha-ras protooncogene product p21, which may be involved in control of cellular growth, is a membrane protein that binds guanine nucleotides and hydrolyzes GTP. p21 GTPase activity is stimulated by lysophosphatidylcholine; a delay in activation was observed unless p21 was incubated with the phospholipid prior to assay. Maximal activation by the phospholipid was observed over a narrow concentration range; the presence in the assay mixture of lysophosphatidylcholine at concentrations above this optimum markedly inhibited p21 GTPase. GTP hydrolysis was also stimulated, but to a lesser degree, by phosphatidylcholine. Phosphatidylinositol and phosphatidylserine did not significantly enhance GTPase activity. The stimulatory effect of phospholipid was mimicked, in part, by nonionic detergents. p21 may be related to other GTPases, the regulatory guanine nucleotide-binding G proteins of the hormone-sensitive adenylate cyclase complex and transducin of the retinal light-activated phosphodiesterase system. The G proteins and transducin are heterotrimers; the alpha subunits possess GTPase activity and the beta gamma subunit complex along with agonist-receptor complex or light-activated rhodopsin enhance GTP hydrolysis. p21 GTPase activity was slightly stimulated by rhodopsin, but, in contrast to the GTPase activity of transducin, stimulation was not light-dependent. GTP hydrolysis was enhanced somewhat by beta gamma subunit complex in the absence, but not in the presence, of rhodopsin. Like the G proteins and transducin, activity of p21 was altered by ADP-ribosylation. Modification of p21 catalyzed by an NAD: arginine ADP-ribosyltransferase purified from turkey erythrocytes decreased both GTPase activity and guanine nucleotide binding activity.
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PMID:Effects of phospholipids and ADP-ribosylation on GTP hydrolysis by Escherichia coli-synthesized Ha-ras-encoded p21. 300 95

Recent studies have shown that the 21-kilodalton protein (p21) Ha-ras gene product shares sequence homology with and may exhibit biochemical properties similar to the mammalian guanine nucleotide-binding proteins. These data suggested that one of the biochemical functions of p21 in the vertebrate cell may be to regulate adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]. We determined both in intact NIH-3T3 murine cells and in membranes isolated from these cells that the hormone-stimulated adenylate cyclase activity of cells expressing the EJ human bladder carcinoma oncogene (EJ-ras) is significantly reduced compared with control cells. Thus, the levels of cAMP measured in the EJ-ras-transformed cells by radioimmunoassay are reduced 78% and 93% after prostaglandin and isoproterenol stimulation, respectively, compared with the levels in control cells. Treatment of the EJ-ras-transformed cells with pertussis toxin or cholera toxin did not correct the alterations in adenylate cyclase activity. Cells expressing the normal human Ha-ras gene displayed intermediate levels of adenylate cyclase hormone sensitivity; these levels of adenylate cyclase activity were greater than those in the EJ-ras-transformed cells but lower than in control cells. Hormone-stimulated adenylate cyclase activities in cells transfected with Rous sarcoma virus DNA were similar to those in control cells. These data support the hypothesis that both the normal and mutated Ha-ras p21s are related to guanine nucleotide-binding proteins.
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PMID:Reduced hormone-stimulated adenylate cyclase activity in NIH-3T3 cells expressing the EJ human bladder ras oncogene. 301 29

Many receptors, in response to ligand activation, trigger inositol phospholipid breakdown, which leads to rapid intracellular responses. The sustained activation of this pathway is believed to be at least one of the factors involved in the stimulation of cell growth and there has been much speculation that certain oncogenes use this pathway to effect uncontrolled cellular proliferation. It has been suggested, by analogy with the receptor-mediated control of adenylate cyclase, that the receptor stimulation of inositol phospholipid metabolism is mediated through a guanine nucleotide regulatory protein (G-protein) called Gp (or Np). Although such a species has not been identified, there is now strong experimental evidence that this process is mediated by a G-protein distinct from the stimulatory and inhibitory G-proteins (Gs and Gi, respectively). The ras genes code for a plasma membrane protein, p21, whose only known biochemical property is a high-affinity GTPase activity. We show here that the expression of normal p21N-ras in NIH 3T3 fibroblasts leads to the coupling of certain growth factor receptors to stimulated inositol phosphate production. We propose that the N-ras proto-oncogene encodes a protein which couples the receptors for certain growth factors to the stimulation of phospholipase C. Thus, N-ras p21 may be the putative Gp or a functionally related protein.
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PMID:Normal p21N-ras couples bombesin and other growth factor receptors to inositol phosphate production. 301 91

The activity of the adenylate cyclase catalytic subunit is higher in Harvey and Kirsten Murine Sarcoma Viruses-infected thyroid epithelial cells than in uninfected control cells either in the presence of Mg2+ alone or following stimulation by Mn2+ or forskolin. The higher activity is associated with an increased cAMP cellular content. The Gpp(NH)p and F- anion are more effective positive modulators in the control than in the virus infected cells: these results exclude therefore that the ras p21 proteins can act as the G-protein alpha-subunit and suggest that they negatively interfere with the G-protein modulation of the adenylate cyclase system.
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PMID:Increased adenylate cyclase activity in rat thyroid epithelial cells expressing viral ras genes. 302 15

In the yeast Saccharomyces cerevisiae, the activation of adenylate cyclase requires the products of the RAS genes and of CDC25. We isolated several dominant extragenic suppressors of the yeast cdc25 mutation. They did not suppress a thermosensitive allele of the adenylate cyclase gene (CDC35). One of these suppressors was a mutated RAS2 gene in which the transition C/G----T/A at position 455 resulted in replacement of threonine 152 by isoleucine in the protein. The same mutation in a v-Ha-ras gene reduces the affinity of p21 for guanine nucleotides (L.A. Feig, B. Pan, T.M. Roberts, and G.M. Cooper, Proc. Natl. Acad. Sci. USA 83:4607-4611, 1986). These results support a model in which the CDC25 gene product is the GDP-GTP exchange factor regulating the activity of the RAS gene product.
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PMID:A new RAS mutation that suppresses the CDC25 gene requirement for growth of Saccharomyces cerevisiae. 304 3

The primary structure of the alpha-subunit of the adenylate cyclase-stimulating G-protein (Gs) has been deduced from the nucleotide sequence of cloned DNA complementary to the bovine cerebral mRNA encoding the polypeptide. Comparison of the amino acid sequences of the alpha-subunits of Gs and transducin reveals that some of the highly conserved regions show sequence homology with elongation factor-Tu and ras p21 proteins and correspond to functional regions of guanine nucleotide-binding proteins.
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PMID:Primary structure of the alpha-subunit of bovine adenylate cyclase-stimulating G-protein deduced from the cDNA sequence. 308 Mar 31

Data indicating that the 21-kDa protein (p21) Harvey-ras gene product shares sequence homology with guanine nucleotide-binding proteins (G proteins) has stimulated research on the influence(s) of p21 on G-protein-regulated systems in vertebrate cells. Our previous work demonstrated that NIH-3T3 mouse cells expressing high levels of the cellular ras oncogene isolated from the EJ human bladder carcinoma (EJ-ras) exhibited reduced hormone-stimulated adenylate cyclase activity. We now report that in these cells another enzyme system thought to be regulated by G proteins is inhibited, namely phospholipases A2 and C. NIH-3T3 cells incubated in plasma-derived serum release significant levels of prostaglandin E2 (PGE2) as determined by radioimmunoassay when exposed to platelet-derived growth factor (PDGF) at 2 units/ml; the levels of PGE2 released from EJ-ras-transfected cells are only 3% those of controls despite a similar basal (unstimulated) release from control and EJ-ras-transfected cells. The lack of PDGF-stimulated PGE2 release from EJ-ras-transfected cells is not due to a defect in the prostaglandin cyclooxygenase enzyme, since incubation of control cells and EJ-ras-transfected cells in 0.33, 3.3, or 33 microM arachidonate resulted in identical levels of PGE2 release. The lack of PDGF-stimulated PGE2 release from EJ-ras-transfected cells also does not result from the loss of functional PDGF receptors. EJ-ras-transformed cells bind 70% as much 125I-labeled PDGF as control cells and are stimulated to incorporate [3H]thymidine and to proliferate after exposure to PDGF. Moreover, this inhibition is not likely the result of a secondary cellular effect related to the transformed phenotype, since NIH-3T3 cells transformed by v-src released PGE2 at wild-type levels after exposure to PDGF. Determination of total water-soluble inositolphospholipids and changes in the specific activities of phosphatidylcholine in control and EJ-ras-transfected cells demonstrated that PDGF-stimulated phospholipase C and A2 activities are inhibited in the EJ-ras-transfected cells.
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PMID:Loss of platelet-derived growth factor-stimulated phospholipase activity in NIH-3T3 cells expressing the EJ-ras oncogene. 309 98

One of the major steps in the understanding of the hormonal and sensory transduction mechanisms in eukaryotic cells has been the discovery of a family of GTP binding proteins which couple receptors to specific cellular effectors. The absolute requirement of GTP for hormonal stimulation of adenylate cyclase was the initial observation which led to the purification of the protein involved: Gs. Gs couples stimulatory receptors to adenylate cyclase. It is a heterotrimer composed of an alpha chain (45 or 52 kDa), a beta chain (35-36 kDa) and a gamma chain (8 kDa). Several other G proteins of known functions have been purified: Gi, which couples inhibitory receptors to adenylate cyclase, and transducin which couples photoexcited rhodopsin to cyclic GMP phosphodiesterase. Some G proteins of uncertain function have also been purified: Go, a G protein mainly localized in nervous tissues and Gp, a G protein isolated from placenta and platelets. All these G proteins have a common design. Like Gs they all consist of 3 chains: alpha, beta and gamma. The beta chains are nearly identical, whereas the gamma chains are more variable. The alpha chains are different, but share common domains (especially at the level of the GTP binding site). These domains of homologies are also similar to those of other GTP binding proteins, such as the product of the ras gene (p21) and the initiation or elongation factors. alpha Chains are also ADP ribosylated by bacterial toxins. Gs and transducin are targets for cholera toxin, whereas Gi, Go and transducin are targets for pertussis toxin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:GTP binding proteins: a key role in cellular communication. 311 13


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