Activation of protein kinase C alters the interaction of α<sub>2</sub>‐adrenoceptors and the inhibitory GTP‐binding protein (G<sub>i</sub>) in human platelets

García‐Sáinz, J. Adolfo; Gutiérrez-Venegas, Gloria

doi:10.1016/0014-5793(89)81588-1

Cited by 18 publications

(10 citation statements)

References 21 publications

(3 reference statements)

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“…Further support for this interpretation was obtained in the binding studies [6]. Our current data are consistent with such an interpretation; they clearly show that the ¢ft'0ct of agents whose receptors activate the GTPase activity of Gi is markedly decreased.…”

Section: Discussionsupporting

confidence: 90%

“…Further support for this interpretation was obtained in binding studies [6]; it was observed that in control platelets, approximately 75% of the receptors were m a high-affinity state for epinephrine and approximately 25°70 in a low-affinity state; Gpp(NH)p shifted the receptor affinity towards the low-affinity conformation [6]. In membranes from TPA-treated platelets, the receptors were in the low-affinity state and no further decrease in affinity was induced by Gpp(NH)p [6]. We examined the effect of TPA on hormone.stimulated GTPase activity; our results indicate that stimulation of PKC markedly diminished the hormone-stimulated GTPase activity of Gi.…”

Section: ! Introductionmentioning

confidence: 64%

“…The findings that activation of PgC in platelets blocks the hormonal inhibition of adenylate cyclase activity [2-61 but not that induced by Gpp(NH)p [5,61 indicated that the rece~tor/Gi interactions rather than the Gi/cyclase interactions were altered [6]. Further support for this interpretation was obtained in the binding studies [6].…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, the ability of hydrolysis-resistant GTP analogues to inhibit adenylate cyclase is not altered by the treatment with "I'PA [5,6] which suggests that the receptor/Gi interaction rather than the Gi/adenylate cyclase interaction is what is altered by PKC. Further support for this interpretation was obtained in binding studies [6]; it was observed that in control platelets, approximately 75% of the receptors were m a high-affinity state for epinephrine and approximately 25°70 in a low-affinity state; Gpp(NH)p shifted the receptor affinity towards the low-affinity conformation [6]. In membranes from TPA-treated platelets, the receptors were in the low-affinity state and no further decrease in affinity was induced by Gpp(NH)p [6].…”

Section: ! Introductionmentioning

confidence: 99%

“…in platelets it has been observed that activation of PKC by 12-tetradecanoyl phorbol 13-acetate (TPA) largely impairs the GTP-dependent hormonesensitive inhibitory pathway to adenylate cyclase which involves the inhibitory GTP-binding protein, Gi [2][3][4][5][6]. It has been shown that PKC phosphorylates Gi [3] and blocks its function in hormonal inhibition of adenylate cyclase.…”

Section: ! Introductionmentioning

confidence: 99%

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Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Gutiérrez-Venegas

García‐Sáinz

1991

FEBS Letters

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The elTwt of 12,tetradecanoyl phorbol 13.n~.etate on the GTPase activity of Gi was invextil|ated. Treatment with TPA did not alter basal GTPa=e activity of membrane= or |he stimulalory effect or prosta~,landin E= (ptltatiwly via Gs), In contrast, the phorbo| c~ter markedly diminished ~timuh. don of GTPase by al~ents who~e receptor= are coupled to Gt such as epinephrine (:c.adrener=ic action), plntele! a¢tivatintt Etctof or thrombin. Pertus~is toxin catalyzed ADP.ribo~ylation was abe decrca,,ed in membranes from TPA-treated platelets as con'~pared to the ~ontrok, It is sulll|ested th,tt the =Iteration in tile hormonal activation of tile OTPau activity of Oi is ~¢ondary to a perturbation in the receptor.Gi interaction.Protei=~ kin=us C; Enzyme activation Gi; GTPase activity; ==.Adreno~cptor !. INTRODUCTION MATERIALS AND METHODSBlood was obtained from healthy men and women wile had taken ~o medication during the previous 2 weeks. Platelet.rich plasma was obtained by centrifuBation. After a preequillbration period of 5 rain at 37"C, the platelets were challenged with I ~,M TPA, or vehicle for I rain; the platelets were centrifuged and homogenized. A crude membrane preparation was obtained as described by Hoffman et zd.GTPase activity was assayed according to Cas~el and $elinger [8], in a mixture containing 2 mM MgCI.,, t m~ 3.isobutyl-l-nl¢th)'lxanthine, O. I mM cyclic AMP, 5 mM creatine phosphate, 1,2 mg/ml ere=line kinase, 0.2% bovine serum albumin, I mM dithiothreitol, 0,1 ntM EDTA, I #M GTP, 50 mM triethanolamine-HCI, pH 7,4° and [~,nPJGTP (0,2 ,~Ci/tube), Tile reaction was initiated by the addition of the membranes (10 #g of protein) and was carried out for l0 rain or the times indicated at 25=C in a final volume of 0.1 ml. The release of [~zP]Pi was determined as described by Aktories and Jakobs [9]. Protein was quantified by the method of Lowry et ~tl.[10]. ADP-ribosylation was assayed in a mixture containing 250 mM potassium phosphate buffer pH 7.5, 10 mM arginine, 5 mM MgCl=, 1 mM A'rP, 10 mM thymidine, 0.'75 mM NADP, 0,1 mM GTP, I0 #M NAP and [nP]NAD (10 ~Ci/tube), The reaction was carried out for 60 min at 30°C for 1 h. Pertussis toxin was activated with 20 rnM DTT for l0 rain at 37°C. The reacti:~, was started by the addition of the membranes (100/.tg); following tt :' reaction, 1 ml of phosphate buffer was added and the membranes were pelleted by centrifugation. The pellet was dissolved and subjected to SDS-PAGE. The gels were fixed, dried and exposed to X.ray film at -72°C [I 1].tmmunoprecipitation of Gi,-, was achieved using a specific polyclonal antibody generated against the decapeptide corresponding to a Gi (KNNLKDCGLF) [13], essentially as described for Gs~ [12]. RESULTSUnder the conditions employed, basal GTPase activity was linear as a function of time of incubation and of similar magnitude in membranes from control and TPA-treated platelets (Fig. 1). Epinephrine (100/~M) and prostaglandin Et (PGEt) (1 #M) increased the rate of GTPase activity (Fig. I) . The effect of epinephrinePublished by E...

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Section: Discussionsupporting

confidence: 90%

Section: ! Introductionmentioning

confidence: 64%

Section: Discussionmentioning

confidence: 99%

Section: ! Introductionmentioning

confidence: 99%

Section: ! Introductionmentioning

confidence: 99%

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Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Gutiérrez-Venegas

García‐Sáinz

1991

FEBS Letters

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Guanine Nucleotide-Binding Proteins in Bipolar Affective Disorder

Manji

Chen²,

Shimon³

et al. 1995

Arch Gen Psychiatry

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Our results are complementary to the previously reported findings of elevated G alpha s levels in postmortem brain tissue form patients with bipolar affective disorder and in mononuclear leukocytes obtained from depressed patients with bipolar (but not unipolar) affective disorder. The significantly higher levels of pertussis toxin-catalyzed [32P]ADP ribosylation in the subjects receiving long-term lithium-treatment replicates our findings in rat cortex and in healthy volunteers and adds to the growing body of evidence implicating G alpha i as a target of lithium's actions.

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Signal Transduction Pathways

Manji

Potter²,

Lenox³

1995

Arch Gen Psychiatry

288

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Lithium remains the most widely used treatment for bipolar disorder, and this monovalent cation represents one of psychiatry's most important treatments. Despite its demonstrated efficacy in reducing both the frequency and severity of recurrent affective episodes and decades of clinical use, the molecular mechanisms underlying its therapeutic actions have not fully been elucidated. In this report, we review the exciting recent progress in the identification of key components of signal transduction pathways (in particular, guanine nucleotide-binding proteins [G proteins], adenylyl cyclases, and protein kinase C isozymes) as targets for lithium's actions and attempt to integrate these effects with the large body of data emphasizing alterations in various neurotransmitter (particularly monoaminergic) systems. Regulation of signal transduction within critical regions of the brain by lithium affects the function of multiple neurotransmitter systems and may thus explain lithium's efficacy in protecting susceptible individuals from spontaneous, stress-induced, and drug-induced cyclic affective episodes. Recent evidence has also demonstrated significant effects of lithium on the regulation of gene expression in the central nervous system, effects that may play a major role in the long-term stabilization of mood. The identification of these intracellular targets for lithium's actions offers the potential for the development of novel, improved therapeutic agents and, in conjunction with molecular genetic approaches, may facilitate our understanding of the biological factors predisposing individuals to manic-depressive illness.

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Activation of protein kinase C alters the interaction of α₂‐adrenoceptors and the inhibitory GTP‐binding protein (G_i) in human platelets

Cited by 18 publications

References 21 publications

Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Guanine Nucleotide-Binding Proteins in Bipolar Affective Disorder

Signal Transduction Pathways

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Activation of protein kinase C alters the interaction of α2‐adrenoceptors and the inhibitory GTP‐binding protein (Gi) in human platelets

Cited by 18 publications

References 21 publications

Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Activation of protein kinase C inhibits hormonal stimulation of the GTPase activity of Gi in human platelets

Guanine Nucleotide-Binding Proteins in Bipolar Affective Disorder

Signal Transduction Pathways

Contact Info

Product

Resources

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Activation of protein kinase C alters the interaction of α₂‐adrenoceptors and the inhibitory GTP‐binding protein (G_i) in human platelets