scholarly journals Regulation of Dynamin I GTPase Activity by G Protein βγ Subunits and Phosphatidylinositol 4,5-Bisphosphate

1996 ◽  
Vol 271 (45) ◽  
pp. 27979-27982 ◽  
Author(s):  
Hsin Chieh Lin ◽  
Alfred G. Gilman
Keyword(s):  
G Protein ◽  
Gtpase Activity ◽  
Dynamin I

scholarly journals Activation of G proteins by guanine nucleotide exchange factors relies on GTPase activity

2015 ◽  
Author(s):  
Rob J Stanley ◽  
Geraint MH Thomas
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Guanine Nucleotide Exchange Factors ◽  
Guanine Nucleotide ◽  
Gtpase Activity ◽  
Nucleotide Exchange ◽  
Signalling Molecules ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factors ◽  
Experimental Strategies

G proteins are an important family of signalling molecules controlled by guanine nucleotide exchange and GTPase activity in what is commonly called an 'activation/inactivation cycle'. The molecular mechanism by which guanine nucleotide exchange factors (GEFs) catalyse the activation of monomeric G proteins is well-established, however the complete reversibility of this mechanism is often overlooked. Here, we use a theoretical approach to prove that GEFs are unable to positively control G protein systems at steady-state in the absence of GTPase activity. Instead, positive regulation of G proteins must be seen as a product of the competition between guanine nucleotide exchange and GTPase activity -- emphasising a central role for GTPase activity beyond merely signal termination. We conclude that a more accurate description of the regulation of G proteins via these processes is as a 'balance/imbalance' mechanism. This result has implications for the understanding of many intracellular signalling processes, and for experimental strategies that rely on modulating G protein systems.

Modulation of Transducin GTPase Activity by Chimeric RGS16 and RGS9 Regulators of G Protein Signaling and the Effector Molecule†

Biochemistry ◽  
1999 ◽  
Vol 38 (16) ◽  
pp. 4931-4937 ◽  
Author(s):  
Randall L. McEntaffer ◽  
Michael Natochin ◽  
Nikolai O. Artemyev
Keyword(s):  
G Protein ◽  
G Protein Signaling ◽  
Gtpase Activity ◽  
Protein Signaling ◽  
Effector Molecule

Calcineurin inhibition of dynamin I GTPase activity coupled to nerve terminal depolarization

Science ◽  
1994 ◽  
Vol 265 (5174) ◽  
pp. 970-973 ◽  
Author(s):  
J. Liu ◽  
A. Sim ◽  
P. Robinson
Keyword(s):  
Nerve Terminal ◽  
Gtpase Activity ◽  
Dynamin I ◽  
Calcineurin Inhibition

Abscisic acid induction of GTP hydrolysis in maize coleoptile plasma membranes

1998 ◽  
Vol 25 (5) ◽  
pp. 539 ◽  
Author(s):  
Helen R. Irving
Keyword(s):  
Abscisic Acid ◽  
G Protein ◽  
G Proteins ◽  
Pertussis Toxin ◽  
Plasma Membranes ◽  
Dependent Manner ◽  
Gtpase Activity ◽  
Heterotrimeric G Proteins ◽  
Gtp Hydrolysis ◽  
Maize Coleoptile

Since receptor-coupled G proteins increase GTP hydrolysis (GTPase) activity upon ligands binding to the receptor, a study was undertaken to determine if abscisic acid (ABA) induced such an effect. Plasma membranes isolated from etiolated maize (Zea mays L.) coleoptiles were enriched in GTPase activity relative to microsomal fractions. Vanadate was included in the assay to inhibit the high levels of vanadate sensitive low affinity GTPases present. Under these conditions, GTPase activity was enhanced by Mg2+, stimulated by mastoparan, and inhibited by GTPγS indicating the presence of either monomeric or heterotrimeric G proteins. The combination of NaF and AlCl3 is expected to inhibit heterotrimeric G protein activity but had little effect on GTPase activity in maize coleoptile membranes. Cholera toxin enhanced basal GTPase activity, confirming the presence of heterotrimeric G proteins in maize plasma membranes. Pertussis toxin also slightly enhanced basal GTPase activity in maize membranes. Abscisic acid enhanced GTPase activity optimally at 5 mmol/L Mg2+ in a concentration dependent manner by 1.5-fold at 10 µmol/L and up to three-fold at 100 µmol/L ABA. Abscisic acid induced GTPase activity was inhibited by GTPγS, the combination of NaF and AlCl3, and pertussis toxin. Overall, these results are typical of a receptor-coupled G protein responding to its ligand.

Pregnancy and estradiol decrease GTPase activity in the guinea pig uterine artery

2001 ◽  
Vol 281 (5) ◽  
pp. H2168-H2175 ◽  
Author(s):  
Irina A. Buhimschi ◽  
Gentzon Hall ◽  
Loren P. Thompson ◽  
Carl P. Weiner
Keyword(s):  
Signal Transduction ◽  
G Protein ◽  
Uterine Artery ◽  
Mesenteric Arteries ◽  
Gtpase Activity ◽  
G Protein Coupling ◽  
High Affinity ◽  
Specific Manner ◽  
Organ Specific ◽  
17Β Estradiol

The mechanisms by which pregnancy redistributes cardiac output in an organ-specific manner are poorly understood. We propose that it is consequential to estrogen-mediated alterations in G protein-mediated signal transduction. Aortas and uterine (UAs) and mesenteric arteries (MAs) were obtained from late-pregnant, nonpregnant, or ovariectomized guinea pigs chronically treated with 17β-estradiol. High-affinity GTPase activity was assayed enzymatically. The cGMP generated in response to the endothelium-dependent agonist ACh was measured in UAs incubated with or without cholera toxin (CTX, which inhibits Gsα). Pregnancy significantly decreased UA but not aorta or MA GTPase activity. 17β-Estradiol decreased UA GTPase activity compared with untreated ovariectomized animals. ACh increased cGMP in pregnant but not nonpregnant UAs. Pretreatment of nonpregnant UAs with CTX increased ACh-induced cGMP levels similar to pregnancy. Thus pregnancy and estradiol decrease the GTPase activity of a CTX-sensitive G protein in UAs, increasing receptor-dependent cGMP release. This alteration in receptor-mediated G protein coupling in UAs may contribute to the characteristic cardiovascular adaptation to pregnancy.

scholarly journals Agonist activation of p42 and p44 mitogen-activated protein kinases following expression of the mouse δ opioid receptor in Rat-1 fibroblasts: effects of receptor expression levels and comparisons with G-protein activation

1996 ◽  
Vol 320 (1) ◽  
pp. 227-235 ◽  
Author(s):  
Andrew R BURT ◽  
I. Craig CARR ◽  
Ian MULLANEY ◽  
Neil G. ANDERSON ◽  
Graeme MILLIGAN
Keyword(s):  
Membrane Protein ◽  
G Protein ◽  
Opioid Receptor ◽  
Gtpase Activity ◽  
Protein Activation ◽  
G Protein Activation ◽  
Mitogen Activated Protein ◽  
Δ Opioid Receptor ◽  
G Protein Coupled ◽  
Stimulation Of

Rat-1 fibroblasts were transfected with a cDNA encoding the mouse Δ opioid receptor. Two separate clones, D2 (which expressed some 6 pmol of the receptor/mg of membrane protein) and DOE (which expressed some 0.2 pmol/mg of membrane protein), were examined in detail. With membranes from both clones, the opioid agonist [D-Ala2]leucine enkephalin (DADLE) caused stimulation of high-affinity GTPase activity and of the binding of guanosine 5´-[γ-[35S]thio]triphosphate, and inhibition of forskolin-amplified adenylate cyclase activity. DADLE also induced phosphorylation and activation of both the p42MAPK (42 kDa isoform) and p44MAPK (44 kDa isoform) members of the mitogen-activated protein kinase (MAP kinase) family. All of these effects of DADLE were prevented in both clones by pretreatment of the cells with pertussis toxin. The maximal response that could be produced by DADLE in direct assays of G-protein activation were substantially greater in clone D2 than in clone DOE, but in both clones essentially full phosphorylation of both p42MAPK and p44MAPK could be achieved. EC50 values for DADLE stimulation of GTPase activity and for activation of p44MAPK were substantially lower in clone D2 than in clone DOE. Moreover, in both clones the EC50 value for DADLE stimulation of p44MAPK was substantially lower than that for stimulation of GTPase activity, and the Hill coefficients for agonist activation of p44MAPK (h > 1) displayed marked co-operativity whereas those for G-protein activation did not (h 0.8–1.0). DADLE activation of p44MAPK showed more sustained kinetics in clone D2 than in clone DOE. By contrast, lysophosphatidic acid, acting at an endogenously expressed G-protein-coupled receptor, also activated p44MAPK in both clones in a pertussis toxin-sensitive manner, but both the kinetics and the concentration–response curve for activation of p44MAPK by this ligand were similar. As with other systems, maintained cellular levels of a cAMP analogue prevented the effects of both G-protein-coupled receptors on activation of p44MAPK. These results demonstrate for the first time that an opioid receptor, at least when expressed in Rat-1 fibroblasts, is able to initiate activation of the MAP kinase cascade in a Gi-dependent manner, and show that only a very small proportion of the cellular Gi population is required to be activated to result in full phosphorylation of the p42MAPK and p44MAPK MAP kinases.

scholarly journals Real-time Detection of Basal and Stimulated G Protein GTPase Activity Using Fluorescent GTP Analogues

2004 ◽  
Vol 280 (9) ◽  
pp. 7712-7719 ◽  
Author(s):  
Emily E. Jameson ◽  
Rebecca A. Roof ◽  
Matthew R. Whorton ◽  
Henry I. Mosberg ◽  
Roger K. Sunahara ◽  
...  
Keyword(s):  
Real Time ◽  
G Protein ◽  
Gtpase Activity ◽  
Real Time Detection

Progesterone, testosterone and estradiol-17β inhibit gonadotropin-releasing hormone stimulation of G protein GTPase activity in plasma membranes from rat anterior pituitary lobe

1992 ◽  
Vol 126 (4) ◽  
pp. 345-349 ◽  
Author(s):  
Rudravajhala Ravindra ◽  
Robert S Aronstam
Keyword(s):  
G Protein ◽  
Anterior Pituitary ◽  
Plasma Membranes ◽  
Gonadotropin Releasing Hormone ◽  
Gonadal Steroids ◽  
Gtpase Activity ◽  
Releasing Hormone ◽  
Low K ◽  
Estradiol 17Β ◽  
Stimulation Of

In order to understand the biochemical mechanisms underlying the rapid, non-genomic effects of gonadal steroids on gonadotropin secretion, we examined the effects of progesterone, testosterone and estradiol-17β on the low Km GTPase activity associated with transducer G proteins coupled to gonadotropin-releasing hormone (GnRH) receptors. Homogenates of anterior pituitary lobes from adult male rats were processed by discontinuous sucrose gradient centrifugation to isolate plasma membranes. The low Km GTPase activity (EC 3.6.1.-) was assayed in 5 μg membrane protein using [γ-32P]GTP at 37°C in an ATP-regenerating buffer containing 1 μmol/l unlabeled GTP. One hundred nmol/l each of progesterone, testosterone and estradiol-17β maximally stimulated low Km GTPase activity by 61%, 59% and 45%, respectively (p<0.05). Time course studies revealed that 100 nmol/l progesterone stimulated the enzyme activity by 93% and 62% at 5 and 30 min, respectively; 100 nmol/l testosterone stimulated GTPase activity by 100% and 72% at 5 and 30 min, respectively: 100 nmol/l estradiol-17β stimulated GTPase activity by 80% and 70% at 5 and 30 min, respectively. GnRH stimulated the low Km GTPase activity by about 60% in a concentration-dependent manner. In the presence of the gonadal steroids, the ability of GnRH to stimulate the GTPase activity was inhibited. For example, stimulation ranged from 36% to 60% with 0.1–100 nmol/l GnRH alone, but only from 7% to 20% in the presence of GnRH and 100 nmol/l progesterone (p<0.05). Similarly, in the presence of 100 nmol/l estradiol-17β, GnRH stimulation of the enzymatic activity ranged from 12% to 19%. It appeared that testosterone was less effective in inhibiting GnRH-stimulated GTPase activity; stimulation ranged from 15% to 32% in the presence of GnRH and 100 nmol/l testosterone (p<0.05). These results, while suggesting that the gonadal steroids disrupt GnRH receptor-G protein interactions, are consistent with the notion that steroids have a profound effect at the membrane level prior to their interaction with the cytosolic receptors.

scholarly journals Trichoderma atroviride G-Protein α-Subunit Gene tga1 Is Involved in Mycoparasitic Coiling and Conidiation

2002 ◽  
Vol 1 (4) ◽  
pp. 594-605 ◽  
Author(s):  
Víctor Rocha-Ramírez ◽  
Carmi Omero ◽  
Ilan Chet ◽  
Benjamin A. Horwitz ◽  
Alfredo Herrera-Estrella
Keyword(s):  
G Protein ◽  
Intracellular Camp ◽  
Protein Kinase Activity ◽  
Gtpase Activity ◽  
Trichoderma Atroviride ◽  
Α Subunit ◽  
Transgenic Lines ◽  
G Protein Α Subunit ◽  
Host Signals ◽  
Camp Levels

ABSTRACT The soil fungus Trichoderma atroviride, a mycoparasite, responds to a number of external stimuli. In the presence of a fungal host, T. atroviride produces hydrolytic enzymes and coils around the host hyphae. In response to light or nutrient depletion, asexual sporulation is induced. In a biomimetic assay, different lectins induce coiling around nylon fibers; coiling in the absence of lectins can be induced by applying cyclic AMP (cAMP) or the heterotrimeric G-protein activator mastoparan. We isolated a T. atroviride G-protein α-subunit (Gα) gene (tga1) belonging to the fungal subfamily with the highest similarity to the Gαi class. Generated transgenic lines that overexpress Gα show very delayed sporulation and coil at a higher frequency. Furthermore, transgenic lines that express an activated mutant protein with no GTPase activity do not sporulate and coil at a higher frequency. Lines that express an antisense version of the gene are hypersporulating and coil at a much lower frequency in the biomimetic assay. The loss of Tga1 in these mutants correlates with the loss of GTPase activity stimulated by the peptide toxin Mas-7. The application of Mas-7 to growing mycelial colonies raises intracellular cAMP levels, suggesting that Tga1 can activate adenylyl cyclase. In contrast, cAMP levels and cAMP-dependent protein kinase activity drop when diffusible host signals are encountered and the mycoparasitism-related genes ech42 and prb1 are highly expressed. Mycoparasitic signaling is unlikely to be a linear pathway from host signals to increased cAMP levels. Our results demonstrate that the product of the tga1 gene is involved in both coiling and conidiation.

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