Diversity of G proteins in Lepidopteran cell lines: Partial sequences of six G protein alpha subunits

2004 ◽  
Vol 57 (3) ◽  
pp. 142-150 ◽  
Author(s):  
Peter J.K. Knight ◽  
Thomas A. Grigliatti
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Cell Lines ◽  
Alpha Subunits ◽  
Lepidopteran Cell Lines

Expression of G-protein alpha subunits in normal rat colonocytes, azoxymethane (AOM)-induced colon tumors, and human colon cancer-derived cell lines

1995 ◽  
Vol 108 (4) ◽  
pp. A954 ◽  
Author(s):  
M.J.G. Bolt ◽  
R.J. Mailloux ◽  
R. Wali ◽  
B. Frawley ◽  
B. Scaglione-Sewell ◽  
...  
Keyword(s):  
Colon Cancer ◽  
G Protein ◽  
Cell Lines ◽  
Human Colon ◽  
Human Colon Cancer ◽  
Colon Tumors ◽  
Alpha Subunits ◽  
Normal Rat

scholarly journals G-proteins in skeletal muscle. Evidence for a 40 kDa pertussis-toxin substrate in purified transverse tubules

1988 ◽  
Vol 254 (2) ◽  
pp. 405-409 ◽  
Author(s):  
M Toutant ◽  
J Barhanin ◽  
J Bockaert ◽  
B Rouot
Keyword(s):  
Skeletal Muscle ◽  
G Protein ◽  
G Proteins ◽  
Pertussis Toxin ◽  
Excitation Contraction Coupling ◽  
Contraction Coupling ◽  
Adp Ribosylation ◽  
Alpha Subunits ◽  
Low Salt ◽  
T Tubules

In muscle, it has been established that guanosine 5′-[gamma-thio]triphosphate (GTP[S]), a non-hydrolysable GTP analogue, elicits a rise in tension in chemically skinned fibres, and that pretreatment with Bordetella pertussis toxin (PTX) decreases GTP[S]-induced tension development [Di Virgilio, Salviati, Pozzan & Volpe (1986) EMBO J. 5, 259-262]. In the present study, G-proteins were analysed by PTX-catalysed ADP-ribosylation and by immunoblotting experiments at cellular and subcellular levels. First, the nature of the G-proteins present in neural and aneural zones of rat diaphragm muscle was investigated. PTX, known to catalyse the ADP-ribosylation of the alpha subunit of several G-proteins, was used to detect G-proteins. Three sequential extractions (low-salt-soluble, detergent-soluble and high-salt-soluble) were performed, and PTX was found to label two substrates of 41 and 40 kDa only in the detergent-soluble fraction. The addition of pure beta gamma subunits of G-proteins to the low-salt-soluble extract did not provide a way to detect PTX-catalysed ADP-ribosylation of G-protein alpha subunits in this hydrophilic fraction. In neural as well as in aneural zones, the 39 kDa PTX substrate, very abundant in the nervous system (Go alpha), was not observed. We then studied the nature of the G alpha subunits present in membranes from transverse tubules (T-tubules) purified from rabbit skeletal muscle. Only one 40 kDa PTX substrate was found in T-tubules, known to be the key element of excitation-contraction coupling. The presence of a G-protein in T-tubule membranes was further confirmed by the immunoreactivity detected with an anti-beta-subunit antiserum. A 40 kDa protein was also detected in T-tubule membranes with an antiserum raised against a purified bovine brain Go alpha. The presence of two PTX substrates (41 and 40 kDa) in equal amounts in total muscle extracts, compared with only one (40 kDa) found in purified T-tubule membranes, suggests that this 40 kDa PTX substrate might be involved in excitation-contraction coupling.

scholarly journals Light-dependent GTP-binding proteins in squid photoreceptors

1990 ◽  
Vol 272 (1) ◽  
pp. 79-85 ◽  
Author(s):  
P R Robinson ◽  
S F Wood ◽  
E Z Szuts ◽  
A Fein ◽  
H E Hamm ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
G Protein ◽  
G Proteins ◽  
Alpha Subunit ◽  
Protein G ◽  
Adp Ribosylation ◽  
Alpha Subunits ◽  
Gtp Binding ◽  
Molecular Masses ◽  
Lines Of Evidence

Previous biochemical and electrophysiological evidence suggests that in invertebrate photoreceptors, a GTP-binding protein (G-protein) mediates the actions of photoactivated rhodopsin in the initial stages of transduction. We find that squid photoreceptors contain more than one protein (molecular masses 38, 42 and 46 kDa) whose ADP-ribosylation by bacterial exotoxins is light-sensitive. Several lines of evidence suggest that these proteins represent distinct alpha subunits of G-proteins. (1) Pertussis toxin and cholera toxin react with distinct subsets of these polypeptides. (2) Only the 42 kDa protein immunoreacts with the monoclonal antibody 4A, raised against the alpha subunit of the G-protein of vertebrate rods [Hamm & Bownds (1984) J. Gen. Physiol. 84. 265-280]. (3) In terms of ADP-ribosylation, the 42 kDa protein is the least labile to freezing. (4) Of the 38 kDa and 42 kDa proteins, the former is preferentially extracted with hypo-osmotic solutions, as demonstrated by the solubility of its ADP-ribosylated state and by the solubility of the light-dependent binding of guanosine 5′-[gamma-thio]triphosphate. The specific target enzymes for the observed G-proteins have not been established.

scholarly journals Agonist activation of transfected human M1 muscarinic acetylcholine receptors in CHO cells results in down-regulation of both the receptor and the α subunit of the G-protein Gq

1993 ◽  
Vol 289 (1) ◽  
pp. 125-131 ◽  
Author(s):  
I Mullaney ◽  
M W Dodd ◽  
N Buckley ◽  
G Milligan
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Cho Cells ◽  
Inositol Phosphate ◽  
Muscarinic Acetylcholine Receptors ◽  
Down Regulation ◽  
Α Subunit ◽  
Muscarinic Acetylcholine ◽  
Alpha Subunits ◽  
M1 Muscarinic

CHO cells stably transfected with cDNA encoding the human M1 muscarinic acetylcholine (HM1) receptor were treated with the cholinergic agonist carbachol at various concentrations for differing times. Levels of the HM1 receptor and of a range of G-proteins were subsequently measured. Carbachol treatment of the transfected cells caused a substantial down-regulation of cellular levels of the alpha subunit of Gq (Gq alpha), but did not significantly alter cellular levels of the alpha subunits of Gs or Gi2. A small decrease in levels of G-protein beta-subunit was also produced. Parallel assessment of agonist-induced down-regulation of the HM1 receptor demonstrated that it was lost in concert with the G-protein. Similar concentrations of carbachol (5 microM) were required to produce half-maximal stimulation of inositol phosphate generation and loss of each of the HM1 receptor and Gq alpha, and half-maximal losses of both receptor and Gq alpha were produced by 3 h of treatment with 1 mM-carbachol. By contrast, treatment of the non-transfected parental CHO cells, which do not express detectable levels of the receptor, with carbachol had no effect on cellular Gq alpha levels. Concurrent treatment of the HM1-expressing CHO cells with carbachol and cycloheximide indicated that suppression of protein synthesis de novo did not mimic the effect of carbachol, and hence even complete inhibition of transcription of the Gq alpha gene and/or translation of pre-existing Gq alpha mRNA could not account for the agonist-induced effect. We have previously noted that cellular levels of both Gs alpha [McKenzie and Milligan (1990) J. Biol. Chem. 265, 17084-17093] and the alpha subunits of the pertussis-toxin-sensitive G-proteins Gi1, Gi2 and Gi3 [Green, Johnson and Milligan (1990) J. Biol. Chem. 265, 5206-5210] can be regulated in certain cell systems by agonist activation of receptors expected to interact with these G-proteins. These results demonstrate that the same is true of Gq alpha and suggest that agonist-induced co-ordinate loss of receptors and associated G-proteins may be a more common feature than has been appreciated to date.

scholarly journals Homologous and unique G protein alpha subunits in the nematode Caenorhabditis elegans.

1991 ◽  
Vol 2 (2) ◽  
pp. 135-154 ◽  
Author(s):  
M A Lochrie ◽  
J E Mendel ◽  
P W Sternberg ◽  
M I Simon
Keyword(s):  
Amino Acid ◽  
G Protein ◽  
G Proteins ◽  
Amino Acid Sequences ◽  
Alpha Subunit ◽  
Predicted Amino Acid Sequence ◽  
C Elegans ◽  
Alpha Subunits ◽  
G Protein Alpha Subunit ◽  
Protein Alpha Subunit

A cDNA corresponding to a known G protein alpha subunit, the alpha subunit of Go (Go alpha), was isolated and sequenced. The predicted amino acid sequence of C. elegans Go alpha is 80-87% identical to other Go alpha sequences. An mRNA that hybridizes to the C. elegans Go alpha cDNA can be detected on Northern blots. A C. elegans protein that crossreacts with antibovine Go alpha antibody can be detected on immunoblots. A cosmid clone containing the C. elegans Go alpha gene (goa-1) was isolated and mapped to chromosome I. The genomic fragments of three other C. elegans G protein alpha subunit genes (gpa-1, gpa-2, and gpa-3) have been isolated using the polymerase chain reaction. The corresponding cosmid clones were isolated and mapped to disperse locations on chromosome V. The sequences of two of the genes, gpa-1 and gpa-3, were determined. The predicted amino acid sequences of gpa-1 and gpa-3 are only 48% identical to each other. Therefore, they are likely to have distinct functions. In addition they are not homologous enough to G protein alpha subunits in other organisms to be classified. Thus C. elegans has G proteins that are identifiable homologues of mammalian G proteins as well as G proteins that appear to be unique to C. elegans. Study of identifiable G proteins in C. elegans may result in a further understanding of their function in other organisms, whereas study of the novel G proteins may provide an understanding of unique aspects of nematode physiology.

Restricted spatial and temporal expression of G-protein alpha subunits during Drosophila embryogenesis

Development ◽  
1991 ◽  
Vol 113 (2) ◽  
pp. 527-538 ◽  
Author(s):  
W.J. Wolfgang ◽  
F. Quan ◽  
N. Thambi ◽  
M. Forte
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Sensory Transduction ◽  
Temporal Expression ◽  
Common Component ◽  
Signaling Systems ◽  
Alpha Subunits ◽  
Drosophila Embryogenesis ◽  
G Protein Coupled

Of the known signal transduction mechanisms, the most evolutionarily ancient is mediated by a family of heterotrimeric guanine nucleotide binding proteins or G proteins. In simple organisms, this form of sensory transduction is used exclusively to convey signals of developmental consequence. In metazoan organisms, however, the developmental role of G-protein-coupled sensory transduction has been more difficult to elucidate because of the wide variety of signals (peptides, small molecules, odorants, hormones, etc.) that use this form of sensory transduction. We have begun to examine the role of G-protein-coupled signaling during development by investigating the expression during Drosophila embryogenesis of a limited set of G proteins. Since these proteins are a common component of all G-protein-coupled signaling systems, their developmental pattern of expression should indicate when and where programmed changes in gene activity are initiated by, or involve the participation of, G-protein-coupled signaling events. We have focused on the spatial and temporal expression pattern of three different Drosophila G-protein alpha subunits by northern blot analysis, in situ hybridization and immunocytochemistry using antibodies directed to peptides specifically found in each alpha subunit. From the spatial and temporal restriction of the expression of each protein, our results suggest that different forms of G-protein-coupled sensory transduction may mediate developmental interactions during both early and late stages of embryogenesis and may participate in a variety of specific developmental processes such as the establishment of embryonic position, the ontogeny of the nervous system and organogenesis.

Expression and potential functions of G-protein alpha subunits in embryos of Xenopus laevis

Development ◽  
1992 ◽  
Vol 116 (1) ◽  
pp. 141-146 ◽  
Author(s):  
A.P. Otte ◽  
L.L. McGrew ◽  
J. Olate ◽  
N.M. Nathanson ◽  
R.T. Moon
Keyword(s):  
Signal Transduction ◽  
G Protein ◽  
G Proteins ◽  
Expression Patterns ◽  
Neural Induction ◽  
Neural Tissue ◽  
Signal Transduction Pathways ◽  
Alpha Subunits ◽  
Whole Mount

During early embryonic development, many inductive interactions between tissues depend on signal transduction processes. We began to test the possibility that G-proteins participate in the signal transduction pathways that mediate neural induction. The expression during Xenopus development of three G alpha subunits, G alpha 0, G alpha i-1 and G alpha s-1, was characterized. The three maternally expressed genes showed different expression patterns during early development. Whole-mount in situ hybridization revealed that all three genes were expressed almost exclusively in the gastrula ectoderm and predominantly in the neuroectoderm in the neurula embryo. In order to investigate the involvement of these proteins in neural induction, we overexpressed the G-protein alpha subunits by injecting the G alpha mRNAs into fertilized eggs. Overexpression of G alpha s-1 increased the ability of gastrula ectoderm to become induced to neural tissue approximately four-fold. Overexpression of G alpha 0 and G alpha i-1 had less pronounced effects on neural competence, and inhibition of the G alpha 0 and G alpha i-1 proteins by pertussis toxin did not change the neural competence of the exposed gastrula ectoderm. Overexpression of the G alpha 0 and G alpha i-1 genes did, however, inhibit the normal disappearance of the blastocoel during gastrulation, suggesting a role for these G-proteins in regulating this process. The data also suggest a specific role for the G alpha s subunit in mediating the initial phases of neural induction.

Presence of P2X1 Purinoceptors in Human Platelets and Megakaryoblastic Cell Lines

1997 ◽  
Vol 78 (06) ◽  
pp. 1500-1504 ◽  
Author(s):  
Catherine Vial ◽  
Béatrice Hechier ◽  
Catherine Léon ◽  
Jean-Pierre Cazenave ◽  
Christian Gachet
Keyword(s):  
Intracellular Calcium ◽  
G Proteins ◽  
Cell Lines ◽  
Calcium Influx ◽  
Human Platelets ◽  
P2y1 Receptor ◽  
Calcium Response ◽  
Ionotropic Receptors ◽  
External Calcium

SummaryHuman platelets are thought to possess at least two subtypes of purinoceptor, one of which, coupled to G-proteins, could be the P2Y1 receptor (Léon et al. 1997). However, it has been suggested that the unique rapid calcium influx induced by ADP in platelets could involve P2X1 ionotropic receptors (MacKenzie et al. 1996) and the aim of this study was thus to investigate the presence of P2X purinoceptors in platelets and megakaryoblastic cells. Using PCR experiments, we found P2X1 mRNA to be present in human platelets and megakaryoblastic cell lines. In platelets, the selective P2X1 agonist αβMeATP induced a rise in intracellular calcium only in the presence of external calcium and this effect was antagonized by suramin and PPADS. Repeated addition of a�MeATP desensitized the P2X1 purinoceptor but only slightly affected the ADP response, while no calcium response to αβMeATP was observed in megakaryoblastic cells. These results support the existence of functional P2X1 purinoceptors on human platelets and the presence of P2X1 transcripts in megakaryoblastic cell lines.

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