Mastoparan, a peptide toxin from wasp venom, mimics receptors by activating GTP-binding regulatory proteins (G proteins).

This study evaluates the involvement of GTP-dependent regulatory proteins (G-proteins) in the regulation of Na+-K+-ATPase activity in proximal convoluted tubule (PCT) segments. Single PCT segments were dissected from rat kidney and permeabilized to allow nucleotides and medium free access to the interior of the cell. A GDP analogue that blocks GTP-dependent activation of the G-protein, GDP beta S (400 microM) significantly inhibited PCT Na+-K+-ATPase activity when Na in the medium (Nam) was greater than or equal to 70 mM. The inhibition was attenuated when Nam was 55 and 35 mM and was no longer significant when Nam was 25 mM. GDP beta S had no inhibitory effect on the activity of purified Na+-K+-ATPase. A nonhydrolyzable GTP analogue, GppNHp (50 microM) significantly increased Na+-K+-ATPase activity when Nam was 25 and 35 mM, but not when Nam was 55-140 mM. Dopamine (DA) and DA1 plus DA2 agonists significantly inhibit Na+-K+-ATPase activity. DA inhibition was competitively abolished by GppNHp. In PCT segments from rats pretreated with pertussis toxin, DA and DA1 plus DA2 agonist inhibition of Na+-K+-ATPase activity was abolished. In PCT segments from rats pretreated with cholera toxin, basal Na+-K+-ATPase activity was increased, but DA significantly inhibited Na+-K+-ATPase activity. Na+-K+-ATPase activity in PCT segments is regulated via a G-protein that stimulates Na+-K+-ATPase activity and a DA-activated pertussis toxin-sensitive G-protein that inhibits Na+-K+-ATPase activity.(ABSTRACT TRUNCATED AT 250 WORDS)

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A slowly ADP-ribosylated pertussis-toxin-sensitive GTP-binding regulatory protein is required for vasopressin-stimulated Ca2+ inflow in hepatocytes

Biochemical Journal ◽

10.1042/bj2990399 ◽

1994 ◽

Vol 299 (2) ◽

pp. 399-407 ◽

Cited By ~ 17

Author(s):

L A Berven ◽

B P Hughes ◽

G J Barritt

Keyword(s):

G Protein ◽

G Proteins ◽

Pertussis Toxin ◽

Single Cells ◽

Regulatory Protein ◽

Regulatory Proteins ◽

Inositol Polyphosphates ◽

Gtp Binding ◽

Inositol 1,4,5 Trisphosphate ◽

Epidermal Growth

The roles of heterotrimeric GTP-binding regulatory proteins (G-proteins) and inositol polyphosphates in the mechanism by which vasopressin stimulates Ca2+ inflow in hepatocytes were investigated by using single cells loaded with fura2 by microinjection. Vasopressin-stimulated Ca2+ inflow was mimicked by microinjection of guanosine 5′-[gamma-thio]triphosphate (GTP[S]) or guanosine 5′-[beta gamma-imido]triphosphate to the cells, but not adenosine 5′-[gamma-thio]triphosphate (ATP[S]) or guanosine 5′-[beta-thio]diphosphate (GDP[S]). Extracellular Gd3+ (5 microM) inhibited both vasopressin- and GTP[S]-stimulated Ca2+ inflow. GDP[S], but not GMP, administered to hepatocytes by microinjection, completely inhibited vasopressin-stimulated Ca2+ inflow and partially inhibited vasopressin-induced release of Ca2+ from intracellular stores. The microinjection of pertussis toxin had no effect either on the release of Ca2+ from intracellular stores or on Ca2+ inflow induced by vasopressin, but completely inhibited changes in these processes induced by epidermal growth factor (EGF). Hepatocytes isolated from rats treated with pertussis toxin for 24 h exhibited no vasopressin- or GTP[S]-stimulated Ca2+ inflow, whereas the vasopressin-stimulated release of Ca2+ from intracellular stores was similar to that observed for control cells. Heparin or ATP[S] inhibited, or delayed the onset of, both vasopressin-induced release of Ca2+ from intracellular stores and vasopressin-stimulated Ca2+ inflow. Vasopressin-induced oscillations in intracellular [Ca2+] were observed in some heparin-treated cells. It is concluded that the stimulation by vasopressin of Ca2+ inflow to hepatocytes requires inositol 1,4,5-trisphosphate (InsP3) and, by implication, the pertussis-toxin-insensitive G-protein required for the activation of phospholipase C beta [Taylor, Chae, Rhee and Exton (1991) Nature (London) 350, 516-518], and another G-protein which is slowly ADP-ribosylated by pertussis toxin and acts between InsP3 and the putative plasma-membrane Ca2+ channel. EGF-stimulated Ca2+ inflow involves at least one G-protein which is rapidly ADP-ribosylated and is most likely required for InsP3 formation.

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Direct activation of GTP-binding regulatory proteins (G-proteins) by substance P and compound 48/80

FEBS Letters ◽

10.1016/0014-5793(90)80023-c ◽

1990 ◽

Vol 259 (2) ◽

pp. 260-262 ◽

Cited By ~ 239

Author(s):

M. Mousli ◽

C. Bronner ◽

Y. Landry ◽

J. Bockaert ◽

B. Rouot

Keyword(s):

Substance P ◽

G Proteins ◽

Regulatory Proteins ◽

Direct Activation ◽

Gtp Binding

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Characterization of GTP-binding proteins in Golgi-associated membrane vesicles from rat adipocytes

Biochemical Journal ◽

10.1042/bj2830795 ◽

1992 ◽

Vol 283 (3) ◽

pp. 795-801 ◽

Cited By ~ 12

Author(s):

A Schürmann ◽

W Rosenthal ◽

G Schultz ◽

H G Joost

Keyword(s):

G Proteins ◽

Glucose Transport ◽

Subcellular Distribution ◽

Sucrose Gradient ◽

Binding Proteins ◽

Glucose Transporters ◽

Beta Subunits ◽

Transport Activity ◽

Gtp Binding Proteins ◽

Gtp Binding

We have previously reported that guanine nucleotides inhibit glucose transport activity reconstituted from adipocyte membrane fractions. In order to further investigate the hypothetical involvement of guanine-nucleotide-binding proteins (GTP-binding proteins) in the regulation of insulin-sensitive glucose transport activity, we studied their subcellular distribution in adipocytes treated or not with insulin. Adipocytes were homogenized and fractionated to yield plasma membranes (PM) and a Golgi-enriched fraction of intracellular membranes (low-density microsomes, LDM). In these membrane fractions, total guanosine 5′-[gamma-[35S]thio]triphosphate ([35S]GTP[S]) binding, alpha- and beta-subunits of heterotrimeric G-proteins, proto-oncogenes Ha-ras and K-ras, and 23-28 kDa GTP-binding proteins were assayed. The levels of alpha s and alpha i (the alpha-subunits of Gs and Gi) were approx. 8-fold lower in LDM than in PM; beta-subunits, Ha-ras and K-ras were not detectable in LDM. Total GTP[S]-binding sites and 23-28 kDa GTP-binding proteins were present in LDM in approximately the same concentrations as in PM. Insulin gave rise to the characteristic translocation of glucose transporters, but failed to alter the subcellular distribution of any of the GTP-binding proteins. Fractionation of the LDM on a discontinuous sucrose gradient revealed that alpha s and alpha i, as detected with antiserum against a common peptide sequence (alpha common), and the bulk of the 23-28 kDa G-proteins sedimented at different sucrose densities. None of the GTP-binding proteins co-sedimented with glucose transporters. Furthermore, the inhibitory effect of GTP[S] on the reconstituted transport activity was lost in the peak fractions of glucose transporters partially purified on the sucrose gradient. These data indicate that LDM from adipocytes contain several GTP-binding proteins in discrete vesicle populations. However, the intracellular GTP-binding proteins are not tightly associated with the vesicles containing the glucose transporter.

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Erratum

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.1991.126 ◽

1991 ◽

Vol 11 (4) ◽

pp. 706-706

Keyword(s):

Blood Flow ◽

Cerebral Blood Flow ◽

Equilibrium State ◽

Rat Brain ◽

G Proteins ◽

Pertussis Toxin ◽

Adp Ribosylation ◽

Gtp Binding ◽

Reconstituted System ◽

Α Subunits

Ischemia of Rat Brain Decreases Pertussis Toxin-Catalyzed [32P] ADP Ribosylation of GTP-Binding Proteins (Gi1 and G0) in Membranes Katsunobu Takenaka, Yasunori Kanaho, Koh-ichi Nagata, Noboru Sakai, Hiromu Yamada, Yoshinori Nozawa [ Originally published in Journal of Cerebral Blood Flow and Metabolism 1991;11:155–160] On page 158 of the above, arrows were erroneously deleted from the equation in the following passage: Heterotrimers of G proteins that bind GDP to α subunits seem to be the preferred substrates for PTcatalyzed ADP ribosylation since guanine nucleotides (GDP and GTP) and 13'Y subunits stimulate ADP ribosylation in the reconstituted system and in membranes (Tsai et aI., 1984). These results indicate that the G proteins may exist at the equilibrium state as shown below: This omission was the result of a typesetting error, which the publisher regrets.

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