Heterotrimeric guanosine triphosphate-binding protein-coupled modulatory actions of motilin on K+channels and postsynaptic γ-aminobutyric acid receptors in mouse medial vestibular nuclear neurons

Abstract Background: It has been shown that γ-aminobutyric acid exerts excitatory actions on the immature brain due to the increased expression of Na+–K+–2Cl− cotransporter isoform 1. The authors sought to clarify whether midazolam, a γ-aminobutyric acid–mimetic hypnotic agent, causes neuronal excitation that can be blocked by bumetanide, a selective inhibitor of Na+–K+–2Cl− cotransporter isoform 1. Furthermore, the authors examined whether bumetanide potentiates the sedative effects of midazolam in neonatal rats. Methods: The authors measured the effects of midazolam with or without bumetanide on the cytosolic Ca2+ concentration ([Ca]2+i) in hippocampal slices (n = 3 in each condition) from rats at postnatal days 4, 7, and 28 (P4, P7, and P28) using fura-2 microfluorometry. Neuronal activity in the hippocampus and thalamus after intraperitoneal administration of midazolam with or without bumetanide was estimated by immunostaining of phosphorylated cyclic adenosine monophosphate–response element–binding protein (n = 12 in each condition). Furthermore, the authors assessed effects of bumetanide on the sedative effect of midazolam by measuring righting reflex latency (n = 6 in each condition). Results: Midazolam significantly increased [Ca]2+i in the CA3 area at P4 and P7 but not at P28. Bumetanide inhibited midazolam-induced increase in [Ca]2+i. Midazolam significantly up-regulated phosphorylated cyclic adenosine monophosphate–response element–binding protein expression in a bumetanide-sensitive manner in the hippocampus at P7 but not P28. Bumetanide enhanced the sedative effects of midazolam in P4 and P7 but not P28 rats. Conclusion: These results suggest that γ-aminobutyric acid A receptor–mediated excitation plays an important role in attenuated sedative effects of midazolam in immature rats.

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The Docking Stage of Yeast Vacuole Fusion Requires the Transfer of Proteins from a Cis-Snare Complex to a Rab/Ypt Protein

The Journal of Cell Biology ◽

10.1083/jcb.148.6.1231 ◽

2000 ◽

Vol 148 (6) ◽

pp. 1231-1238 ◽

Cited By ~ 150

Author(s):

Albert Price ◽

Darren Seals ◽

William Wickner ◽

Christian Ungermann

Keyword(s):

Binding Protein ◽

Snare Complex ◽

Guanosine Triphosphate ◽

Yeast Vacuole ◽

Vacuole Fusion ◽

Direct Association ◽

Rab Family

The homotypic fusion of yeast vacuoles requires Sec18p (NSF)-driven priming to allow vacuole docking, but the mechanism that links priming and docking is unknown. We find that a large multisubunit protein called the Vam2/6p complex is bound to cis-paired SNAP receptors (SNAREs) on isolated vacuoles. This association of the Vam2/6p complex with the cis-SNARE complex is disrupted during priming. The Vam2/6p complex then binds to Ypt7p, a guanosine triphosphate binding protein of the Rab family, to initiate productive contact between vacuoles. Thus, cis-SNARE complexes can contain Rab/Ypt effectors, and these effectors can be mobilized by NSF/Sec18p-driven priming, allowing their direct association with a Rab/Ypt protein to activate docking.

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Role of pertussis toxin-sensitive guanosine triphosphate-binding proteins in the response of erythroblasts to erythropoietin

Blood ◽

10.1182/blood.v77.3.486.486 ◽

1991 ◽

Vol 77 (3) ◽

pp. 486-492 ◽

Cited By ~ 16

Author(s):

BA Miller ◽

K Foster ◽

JD Robishaw ◽

CF Whitfield ◽

L Bell ◽

...

Keyword(s):

Adenylate Cyclase ◽

Pertussis Toxin ◽

Binding Proteins ◽

Binding Protein ◽

Cyclase Activity ◽

Adenylate Cyclase Activity ◽

Guanosine Triphosphate ◽

Gtp Binding Protein ◽

Gtp Binding

Abstract Human progenitor-derived erythroblasts have been recently shown to respond to erythropoietin (Epo) with an increase in intracellular free calcium concentration [Cac]. To explore the role of guanosine triphosphate (GTP)-binding proteins in mediating the rise in [Cac], single day 10 erythroid burst forming unit (BFU-E)-derived erythroblasts loaded with Fura-2 were pretreated with pertussis toxin (PT), stimulated with Epo, and [Cac] measured over 18 minutes with fluorescence microscopy coupled to digital video imaging. The [Cac] increase in day 10 erythroblasts stimulated with Epo was blocked by pretreatment with PT in a dose-dependent manner but not by heat- inactivated PT. These observations provided strong evidence that a PT- sensitive GTP-binding protein is involved. To further characterize the GTP-binding protein, day 10 erythroblast membrane preparations were solubilized, electrophoresed, and immunoblotted with antibodies specific for the known PT-sensitive G-protein subunits: the three subtypes of Gia (1,2, and 3) and Goa, Gia1 or Gia3 and Gia2 were identified but no Goa was found. To examine the influence of Epo on adenylate cyclase activity, day 10 erythroblasts were initially treated with Epo, isolated membrane preparations made, and cyclic adenosine monophosphate (cAMP) production by adenylate cyclase in membrane preparations in the presence of theophylline measured. Epo did not inhibit but significantly stimulated adenylate cyclase activity. However, the mechanism of increase of [Cac] appears to be independent of adenylate cyclase stimulation because treatment of erythroblasts with the cell-permeant dibutyryl cAMP failed to increase [Cac]. In summary, pertussis toxin blocks the increase in [Cac] in erythroblasts after Epo stimulation suggesting that this response is mediated through a pertussis toxin-sensitive GTP-binding protein. Candidate PT-sensitive GTP-binding proteins identified on day 10 erythroblasts were Gia 1, 2, or 3, but not Goa.

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Epoxyeicosatrienoic Acids Activate K+Channels in Coronary Smooth Muscle Through a Guanine Nucleotide Binding Protein

Circulation Research ◽

10.1161/01.res.80.6.877 ◽

1997 ◽

Vol 80 (6) ◽

pp. 877-884 ◽

Cited By ~ 179

Author(s):

Pin-Lan Li ◽

William B. Campbell

Keyword(s):

Smooth Muscle ◽

Binding Protein ◽

Nucleotide Binding ◽

Epoxyeicosatrienoic Acids ◽

Guanine Nucleotide ◽

Guanine Nucleotide Binding Protein ◽

Coronary Smooth Muscle ◽

K Channels ◽

Guanine Nucleotide Binding ◽

Nucleotide Binding Protein

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Variation in the Small Guanosine Triphosphate (GTP)-Binding Protein, Secretion-Associated and RAS-Related (SAR1A) Gene and Response to Hydroxyurea Treatment in Sickle Cell Disease.

Blood ◽

10.1182/blood.v110.11.3392.3392 ◽

2007 ◽

Vol 110 (11) ◽

pp. 3392-3392

Author(s):

Chutima Kumkhaek ◽

Jianqiong Zhu ◽

James G. Taylor ◽

Carolyn Hoppe ◽

Gregory J. Kato ◽

...

Keyword(s):

Sickle Cell Disease ◽

Confidence Interval ◽

Sickle Cell ◽

Protein Secretion ◽

Binding Protein ◽

Erythroid Cell ◽

Cell Disease ◽

Guanosine Triphosphate ◽

Gtp Binding Protein ◽

Gtp Binding

Abstract Hydroxyurea (HU) is a well-known chemotherapeutic agent and a potent ribonuclease reductase inhibitor that induces Hb F synthesis in sickle cell disease and some thalassemia syndromes. Our group has shown that the HU-inducible small guanosine triphosphate (GTP)-binding protein, secretion-associated and RAS-related (SAR) protein plays a role in g-globin gene induction and erythroid cell maturation by causing cell apoptosis and G1/S-phase arrest through the reduction of PI3 kinase and ERK phosphorylation and increased p21 and GATA2 expression. Our preliminary analysis indicates that HU inducibility is mediated at the transcriptional level, and is localized to elements in the SAR1A promoter. The aim of this study was to assess whether polymorphisms in the SAR1A gene promoter are associated with Hb F levels or HU therapeutic responses. We studied 386 sickle cell disease patients consisting of 269 adults treated with or without HU and 117 newborns with sickle cell disease identified from a newborn screening program. Twenty point mutations, including one nonsynonymous variant, were identified in SAR1A, including nine previously reported in SNP databases. A difference in genotype frequencies was observed between adults and newborns for rs2310991 in the 5′UTR (odd ratio [OR] = 1.9, 95% confidence interval [CI] = 1.1–3.2, P=0.009) and +31 T>C in 5′UTR (odd ratio [OR] 9.8 [ 1.3–73.9]; confidence interval [CI] 95%; P<0.001). Three previously unknown SNPs in the upstream 5′UTR (−809 C>T, −502 G>T and −385 C>A) were significantly associated with the HbF response in Hb SS patients treated with HU (P<0.05). Our data suggest that the SAR1A polymorphism might contribute to the regulation of HbF expression and modulate patient responses to HU in sickle cell disease.

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