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L Birnbaumer

Publications and source records attributed to L Birnbaumer.

At least 145 records · Page 8Linked to original sources
PubMed1990

Transduction of receptor signal into modulation of effector activity by G proteins: the first 20 years or so ....

G proteins are central to the transduction of many receptor signals. I review the "firsts" that have led to our current knowledge and delineate some of the issues being investigated in several laboratories worldwide.--Birnbaumer, L. Transudction of receptor signal into modulation of effector activity by G proteins: the first 20 years or so...FASEB J. 4:3068-3078; 1990.

Adenylyl Cyclases↗
PubMed1990

G protein coupling of receptors to ionic channels and other effector systems.

1. Four questions raised by previous studies that had shown activation of K+ channels by alpha subunits of the type 3 Gi protein are addressed in the present communication: a) are K+ channels specific for one Gi? b) are there more ionic channels under direct G protein control? c) can we confirm using recombinant G alpha s the results obtained with biochemically resolved G alpha s and continue ascribing the regulatory effector to this part of the alpha beta gamma holo-G protein? and d) can we confirm that a single G alpha, Gs alpha in this case, is able to affect more than one type of effector function? 2. We found Gi alpha s are isoforms, that there exist also Gi-insensitive, Go-responsive K+ channels and that G alpha s can be multifunctional. Thus, a single receptor will elicit cellular responses that will depend on the endogenous G protein as well as the type of effector function expressed in it. 3. In another set of experiments we found that G beta gamma s, be they derived from human erythrocytes, human placenta, bovine brain or bovine retina, all inhibit Gk-gated K+ channel activity as seen in inside out membrane patches with GTP as the driving nucleotide. In addition we noted that inhibition was much more effective under basal (no agonist in the pipette) than agonist stimulated conditions, as reported in earlier experiments in which beta-adrenoceptors, Gs and catalytic unit of adenylyl cyclase had been incorporated into phospholipid vesicles. 4. We propose that one of the roles of G beta gamma s in membranes is to quench ligand independent G protein activation by unoccupied receptors. Other roles of G beta gamma s are: a) by re-associating with GDP-G alpha s, to promote interaction with receptors, and b) by dissociating from activated R.G alpha *GTP.beta gamma, to allow for receptor dissociation from GTP-activated G alpha s, which is required to satisfy the catalytic mode of receptor action.

GTP-Binding Proteins↗
PubMed1990

Detergents, dimeric G beta gamma, and eicosanoid pathways to muscarinic atrial K+ channels.

Control experiments for the direct effects of G protein beta gamma-subunits (G beta gamma) on muscarinic atrial K+ channel [K+ (ACh)] currents have produced different results (Nature Lond. 327: 21-22, 1987; Nature Lond. 325: 296-297, 1987; Cold Spring Harbor Symp. Quant. Biol. 53: 365-373, 1989). A recent view is that stimulation is indirect via phospholipase by (PLA2) and arachidonic acid (AA) metabolites (Nature Lond. 337: 504-505, 1989). On reexamination we found that 1) the zwitterionic detergent 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate (CHAPS) used to suspend beta gamma stimulates atrial K+ (ACh) currents by itself, and the effects are concentration and Mg2+ dependent; 2) CHAPS stimulates atrial ATP-sensitive K+ channel and inwardly rectifying K+ channel currents; 3) blockers of eicosanoid pathways have nonspecific effects on atrial K+, Ca2+, and Na+ channels. We have confirmed that detergent-free, hydrophilic G beta gamma-subunits inhibit K+ (ACh) currents. Stimulatory effects of dimeric G beta gamma could not be separated from stimulatory effects of detergent, and blockers of PLA2 or lipoxygenase pathways do not clearly establish the significance of these pathways to atrial K+ (ACh) currents.

Animals↗
PubMed1990

Membrane-delimited stimulation of heart cell calcium current by beta-adrenergic signal-transducing Gs protein.

A severalfold increase in calcium current (ICa) is a signal feature of the maximal beta-adrenergic response of the heart. It is generally ascribed to enhanced adenosine 3',5'-cyclic monophosphate (cAMP)-dependent phosphorylation of calcium (Ca) channels after beta-receptor activation of the guanosine nucleotide-binding (G) protein Gs, and Gs activation of the adenylyl cyclase cascade. We blocked phosphorylation pathways in guinea pig cardiomyocytes to unmask other possible ICa-stimulatory modes. In blocked cells, ICa increased by approximately 50% during 1) beta-receptor activation of Gs, 2) intracellular activation of Gs, and 3) intracellular application of preactivated Gs, We conclude that fast, membrane-delimited Gs modulation participates in the physiological regulation of cardiac ICa.

Animals↗
PubMed1990

Coupling of ATP-sensitive K+ channels to A1 receptors by G proteins in rat ventricular myocytes.

ATP-sensitive K+ (K+[ATP]) current is thought to be regulated by GTP-binding proteins (G proteins), but the pathways that couple receptor, G protein, and channel have not been defined. We studied regulation of tolbutamide-sensitive K+[ATP] current in neonatal rat ventricular myocytes. Application of 0.1 mM ATP to the intracellular side of membrane patches reduced K+ [ATP] channel activity, and addition of the nonhydrolyzable GTP analogue guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) at 0.1 mM restored activity. Application of 0.1 mM intracellular GTP plus 10 microM extracellular adenosine or 100 nM N6-cyclohexyladenosine had the same effect as GTP gamma S; hence K+[ATP] channels may be coupled to adenosine receptors via G proteins. To determine which G protein, we applied G alpha subunits, preactivated with GTP gamma S to the cytoplasmic side of membrane patches, and found that alpha i1, alpha i2, and alpha i3 mimicked the effect of GTP gamma S, but not alpha o or Gs, suggesting that Gi alpha acts via a membrane-delimited pathway. Adenosine receptor coupling may be important for activating K+[ATP] channels in ischemic muscle.

Adenosine↗
PubMed1990

G proteins and the mechanism of action of hormones, neurotransmitters, and autocrine and paracrine regulatory factors.

Signal transduction by G proteins is a fundamental and widespread mechanism used by a wide variety of hormones, neurotransmitters, and autocrine and paracrine factors to regulate cellular functions. G proteins modulate not only cAMP formation, but also intracellular Ca2+ mobilization, arachidonic acid release, and, very importantly, membrane potential. The mechanism by which G proteins are activated provides for amplification, reversal of action, and continued monitoring of incoming signals. Not all G proteins are known and some are known but their functions are still unknown. More G proteins and more effector functions affected by them will surely be found. We discuss these features of signal transduction by G proteins.

Arachidonic Acids↗
PubMed1990

Roles of G proteins in coupling of receptors to ionic channels and other effector systems.

Guanine nucleotide binding (G) proteins are heterotrimers that couple a wide range of receptors to ionic channels. The coupling may be indirect, via cytoplasmic agents, or direct, as has been shown for two K+ channels and two Ca2+ channels. One example of direct G protein gating is the atrial muscarinic K+ channel K+[ACh], an inwardly rectifying K+ channel with a slope conductance of 40 pS in symmetrical isotonic K+ solutions and a mean open lifetime of 1.4 ms at potentials between -40 and -100 mV. Another is the clonal GH3 muscarinic or somatostatin K+ channel, also inwardly rectifying but with a slope conductance of 55 pS. A G protein, Gk, purified from human red blood cells (hRBC) activates K+ [ACh] channels at subpicomolar concentrations; its alpha subunit is equipotent. Except for being irreversible, their effects on gating precisely mimic physiological gating produced by muscarinic agonists. The alpha k effects are general and are similar in atria from adult guinea pig, neonatal rat, and chick embryo. The hydrophilic beta gamma from transducin has no effect while hydrophobic beta gamma from brain, hRBCs, or retina has effects at nanomolar concentrations which in our hands cannot be dissociated from detergent effects. An anti-alpha k monoclonal antibody blocks muscarinic activation, supporting the concept that the physiological mediator is the alpha subunit not the beta gamma dimer. The techniques of molecular biology are now being used to specify G protein gating. A "bacterial" alpha i-3 expressed in Escherichia coli using a pT7 expression system mimics the gating produced by hRBC alpha k.

Animals↗
PubMed1990

Roles of G proteins and G protein subunits in signal transduction.

General features of signal transduction by G proteins are described. Recent results on roles of beta gamma dimers, kinetic properties of Gi alpha subunits, structural diversity of Go alpha subunits and possible functions of the Go protein as a stimulator of phospholipase C activity are discussed.

Amino Acid Sequence↗
PubMed1989

Induction of calcium currents by the expression of the alpha 1-subunit of the dihydropyridine receptor from skeletal muscle.

The dihydropyridine (DHP) receptor purified from skeletal muscle comprises five protein subunits (alpha 1, alpha 2, beta, gamma and delta) and produces Ca2+ currents that are blocked by DHPs. Cloning of the alpha 1- and alpha 2-subunits, the former affinity-labelled by DHP, has shown that the alpha 1-subunit is expressed in skeletal muscle alone, whereas the alpha 2- and delta- subunits are also expressed in other tissues. Although the transient expression of the alpha 1-subunit in myoblasts from dysgenic mice (but not in oocytes) has been demonstrated, the use of these expression systems to determine the function of the alpha 1- subunit is complicated by the presence of endogenous Ca2+ currents, which may reflect the constitutive expression of proteins similar to the alpha 2-, beta-, gamma- and/or delta-subunits. We therefore selected a cell line which has no Ca2+ currents or alpha 2- subunit, and probably no delta-subunit for stable transformation with complementary DNA of the alpha 1- subunit. The transformed cells express DHP-sensitive, voltage-gated Ca2+ channels, indicating that the minimum structure of these channels is at most an alpha 1 beta gamma complex and possibly an alpha 1- subunit alone.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1989

Ion channels and G proteins.

Newly discovered relationships offer insight into the mechanisms by which extracellular signals produce complex cellular responses. Ion channels, serving crucial cellular functions, are now known to be gated by G proteins directly (by membrane-delimited processes) and indirectly (by cytoplasmic second messengers). It also appears that a G protein may act on several effectors.

Acetylcholine↗
PubMed1989

Molecular cloning and expression of a fifth muscarinic acetylcholine receptor.

A cDNA of 2149 base pairs with an incomplete open reading frame (ORF) encoding amino acids 1-516 of a 531-amino acid protein highly homologous to muscarinic receptors was cloned from a rat brain cDNA library. The complete ORF was then deduced from a DNA fragment cloned from a rat genomic library. This ORF was subcloned into the eukaryotic expression vector p91023(B) under control of the adenovirus major late promoter and co-transfected with the thymidine kinase selection marker into muscarinic receptor-negative, thymidine kinase-negative murine L cells. Stable transformants were selected and tested for acquisition of muscarinic receptors by following appearance of specific binding sites for the muscarinic ligand [3H] N-methylscopolamine. Two cell lines, LM5.36 and LM5.40, were cloned and shown to express typical muscarinic receptor sites, thus confirming that the newly cloned ORF encodes a muscarinic receptor, the rat M5 muscarinic acetylcholine receptor. Tests for activities showed it to stimulate phosphoinositide hydrolysis in intact cells, without affecting positively or negatively adenylyl cyclase activity. The M5 receptor contains two putative glycosylation sites at its amino terminus and, based on hydropathicity analysis, is predicted to span the plasma membrane seven times. Like 17 other receptors of this class, the M5 receptor has 19 conserved amino acids, among which are 4 prolines located in the 4th, 5th, 6th, and 7th predicted transmembrane regions, conferring possible bends to these helices, and 2 cysteines, one in the 1st and the other in the 2nd extracellular loop, possibly providing for a disulfide bond. Similarity in amino acid composition and in patterns of antagonist binding and biologic effects suggest the M5 receptor to be M1-like.

Adenylyl Cyclases↗
PubMed1989

Molecular cloning and sequence determination of a cDNA coding for the alpha-subunit of a Go-type protein of Xenopus laevis oocytes.

Xenopus laevis oocytes are cells ideally suited to the study of signal transduction and of the G-proteins that are involved in this process. A X. laevis cDNA library in lambda gt10 has been screened with a mixture of three oligonucleotide probes designed to detect sequences found in various mammalian alpha-subunits of G-proteins. One of these clones has been purified through tertiary screening and the DNA insert has been sequenced. This clone was found to include the total sequence coding for a 354 amino acid protein that is 89% identical to the sequence of alpha-subunit of rat Go. The differences with the mammalian protein were clustered in amino acids 290-315, which have been postulated to define the region interacting with the receptor and effector molecule. The homology with the alpha-subunits of other mammalian G-proteins is lower (65-70% to Gi and 42% to Gs). On this basis, this clone can be classified as Go-like.

Amino Acid Sequence↗
PubMed1989

Splice variants of the alpha subunit of the G protein Gs activate both adenylyl cyclase and calcium channels.

Signal transducing guanine nucleotide binding (G) proteins are heterotrimers with different alpha subunits that confer specificity for interactions with receptors and effectors. Eight to ten such G proteins couple a large number of receptors for hormones and neurotransmitters to at least eight different effectors. Although one G protein can interact with several receptors, a given G protein was thought to interact with but one effector. The recent finding that voltage-gated calcium channels are stimulated by purified Gs, which stimulates adenylyl cyclase, challenged this concept. However, purified Gs may have four distinct alpha-subunit polypeptides, produced by alternative splicing of messenger RNA. By using recombinant DNA techniques, three of the splice variants were synthesized in Escherichia coli and each variant was shown to stimulate both adenylyl cyclase and calcium channels. Thus, a single G protein alpha subunit may regulate more than one effector function.

Adenylyl Cyclases↗