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J L Benovic

Publications and source records attributed to J L Benovic.

At least 163 records · Page 9Linked to original sources
PubMed1988

The beta-adrenergic receptor kinase: role in homologous desensitization in S49 lymphoma cells.

Phosphorylation of the beta-adrenergic receptor (beta AR) is closely associated with homologous desensitization of the beta-adrenergic receptor-coupled adenylate cyclase system. Homologous desensitization and receptor phosphorylation also occur in cell mutants which are deficient in their cAMP-dependent protein kinase (kin- mutant of S49 lymphoma cells). beta AR phosphorylation is mediated by a cAMP-independent protein kinase which phosphorylates the receptor only when it is occupied by a beta-agonist. During the time course of desensitization the beta AR kinase (beta ARK) activity is translocated from a cytoplasmic to a plasma membrane location. beta ARK translocation can also be effected by prostaglandin E1 (PGE1) suggesting that this beta ARK may represent a more general enzyme capable of phosphorylating other adenylate cyclase-coupled receptors. Thus, beta ARK may play a key role in the process of homologous desensitization of adenylate cyclase coupled receptors. Extracellular hormones interact with specific receptors at the outer surface of the plasma membrane and thus initiate a cellular response. One of the best studied transmembrane signalling systems known to be coupled to the occupancy of cell surface receptors is adenylate cyclase. The adenylate cyclase system is composed of various components all of which have been purified to homogeneity (Shorr et al., 1982; Homcy et al., 1983; Benovic et al., 1984; Codina et al., 1984; Northup et al., 1980; Sternweis et al., 1981; Bokoch et al., 1984; Pfeuffer et al., 1985). Initially, agonist binding to the receptor promotes coupling of the occupied receptor to one of the guanine nucleotide binding regulatory proteins. These proteins are members of a family of heterotrimeric proteins consisting of alpha, beta and gamma subunits. Stimulatory receptors like the beta-adrenergic (Cerione et al., 1984) or glucagon (Iyengar et al., 1979) receptors couple to the stimulatory regulatory protein Ns (or Gs) whereas inhibitory receptors like the alpha 2-adrenergic (Jacobs et al., 1976) or M2-muscarinic (Harden et al., 1982) receptors couple to the inhibitory regulatory protein Ni (or Gi). Prolonged exposure to agonist hormones, either stimulatory or inhibitory, results in an attenuation of the response to the hormonal activation, a phenomenon called tachyphylaxis or desensitization (Harden, 1983; Sibley and Lefkowitz, 1985; Sharma et al., 1975). One of the best studied models for desensitization is the beta-adrenergic receptor-coupled adenylate cyclase system. In this system two different forms of desensitization have been characterized.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

Cloning of the cDNA and genes for the hamster and human beta 2-adrenergic receptors.

The adenylate cyclase-stimulatory beta 2-adrenergic receptor has been purified to apparent homogeneity from hamster lung. Partial amino acid sequence obtained from isolated CNBr peptides was used to clone the gene and cDNA for this receptor. The predicted amino acid sequence for the hamster beta 2-adrenergic receptor revealed that the protein consists of a single polypeptide chain of 418 aa with consensus N-glycosylation and phosphorylation sites predicted by previous in vitro data. The most striking feature of the receptor protein however, is that it contains seven stretches of hydrophobic residues similar to the proposed seven transmembrane segments of the light receptor rhodopsin. Significant amino acid homology (30-35%) can be found between the hamster beta 2-adrenergic receptor and rhodopsin within these putative membrane spanning regions. Using a hamster beta 2-adrenergic receptor probe, the gene and cDNA for the human beta 2-adrenergic receptor were isolated, revealing a high degree of homology (87%) between the two proteins from different species. Unlike the genes encoding the family of opsin pigments, of which rhodopsin is a member, the genes encoding both hamster and human beta 2-adrenergic receptors are devoid of introns in their coding as well as 5' and 3' untranslated nucleotide sequences. The cloning of the genes and the elucidation of the aa sequences for these G-protein coupled receptors should help to determine the structure-function as well as the evolutionary relationship of these proteins.

Amino Acid Sequence↗
PubMed1987

Agonist-dependent phosphorylation of the alpha 2-adrenergic receptor by the beta-adrenergic receptor kinase.

Desensitization of the beta-adrenergic receptor, a receptor which is coupled to the stimulation of adenylate cyclase, may be regulated via phosphorylation by a unique protein kinase. This recently discovered enzyme, known as the beta-adrenergic receptor kinase, only phosphorylates the agonist-occupied form of the beta-adrenergic receptor. To assess whether receptors coupled to the inhibition of adenylate cyclase might also be substrates, we examined the effects of beta-adrenergic receptor kinase on the partially purified human platelet alpha 2-adrenergic receptor. Phosphorylation of the reconstituted alpha 2-adrenergic receptor was dependent on agonist occupancy and was completely blocked by coincubation with alpha 2-antagonists. The time course of phosphorylation of the alpha 2-adrenergic receptor was virtually identical to that observed with the beta-adrenergic receptor with maximum stoichiometries of 7-8 mol of phosphate/mol of receptor in each case. In contrast, the alpha 1-adrenergic receptor, which is coupled to stimulation of phosphatidylinositol hydrolysis, is not a substrate for the beta-adrenergic receptor kinase. These results suggest that receptors coupled to either stimulation or inhibition of adenylate cyclase may be regulated by an agonist-dependent phosphorylation mediated by the beta-adrenergic receptor kinase.

Affinity Labels↗
PubMed1987

The multiple membrane spanning topography of the beta 2-adrenergic receptor. Localization of the sites of binding, glycosylation, and regulatory phosphorylation by limited proteolysis.

The beta 2-adrenergic receptor (beta-AR) is an integral membrane glycoprotein of apparent Mr approximately equal to 64,000. The amino acid sequence deduced from the beta-AR gene reveals homology with the visual pigment rhodopsin of retinal rod outer segments. We have proposed a structural model of beta-AR which is similar to that elucidated for rhodopsin. In this paper we identify a number of structural and topographical characteristics of beta-AR consistent with the model through the use of limited proteolysis. Limited trypsinization of beta-AR reconstituted in lipid vesicles yields two insoluble (integral membrane) domains of Mr approximately equal to 38,000 and 26,000. Identical results were obtained in intact cells, indicating that the cleavage site of the receptor is accessible at the extracellular surface of the plasma membrane. The amino-terminal domain (38 kDa) contains the ligand binding site (as revealed by photoaffinity labeling) and the sites of glycosylation (as revealed by its sensitivity to endoglycosidase F), whereas the carboxyl-terminal domain (26 kDa) contains all the sites of in vitro phosphorylation by cAMP-dependent protein kinase and the beta-adrenergic receptor kinase. Of four canonical sites for N-linked glycosylation, two near the amino and two near the carboxyl terminus, only those in the amino-terminal domain (Asn6 and Asn15) are utilized and sensitive to endoglycosidase F. Carboxypeptidase Y treatment of reconstituted native beta-adrenergic receptor generates a truncated (approximately 57 kDa) glycopeptide that has lost most of the sites phosphorylated by beta-AR kinase and one of the sites phosphorylated by protein kinase A. The various features delineated, including the length of the carboxypeptidase Y-sensitive region, the extracellular location of the trypsin-sensitive site, the location of the sites of phosphorylation and glycosylation all constrain the receptor to a rhodopsin-like structure with multiple membrane spanning segments.

Amino Acid Sequence↗
PubMed1987

Purification and characterization of the beta-adrenergic receptor kinase.

The beta-adrenergic receptor kinase (beta-ARK) is a recently discovered enzyme which specifically phosphorylates the agonist-occupied form of the beta-adrenergic receptor (beta-AR) as well as the light-bleached form of rhodopsin. beta-ARK is present in a wide variety of mammalian tissues. The kinase can be purified from bovine cerebral cortex to greater than 90% homogeneity by sequential chromatography on Ultrogel AcA34, DEAE-Sephacel, CM-Fractogel, and hydroxylapatite. This results in an approximately 20,000-fold purification with an overall recovery of 12%. The purified kinase has an Mr approximately 80,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Several findings indicate that this peptide contains the beta-ARK activity. First, on hydroxylapatite chromatography the enzyme activity coelutes with the Mr approximately 80,000 protein as revealed by Coomassie-Blue staining. Second, under phosphorylating conditions the Mr approximately 80,000 protein is phosphorylated. Finally, the Mr approximately 80,000 protein specifically interacts with reconstituted agonist-occupied beta-AR. Kinetic parameters of the enzyme for beta-AR are Km = 0.25 microM and Vmax = 78 nmol/min/mg whereas for rhodopsin the values are Km = 6 microM and Vmax = 72 nmol/min/mg. The Km value of the enzyme for ATP is approximately 35 microM using either beta-AR or rhodopsin as substrate. Receptor phosphorylation by beta-ARK is effectively inhibited by Zn2+, digitonin and a variety of salts. The availability of purified beta-ARK should greatly facilitate studies of its role in receptor desensitization.

Animals↗
PubMed1987

Somatostatin induces translocation of the beta-adrenergic receptor kinase and desensitizes somatostatin receptors in S49 lymphoma cells.

The beta-adrenergic receptor kinase is a cytosolic enzyme that specifically phosphorylates the agonist-occupied form of the beta-adrenergic receptor (beta AR). Beta AR kinase appears to be translocated from the cytosol to the plasma membrane when kin- S49 lymphoma cells are incubated with either beta-adrenergic agonists or prostaglandin E1, both of which act through receptors which stimulate adenylate cyclase. We report here that brief (approximately 20 min) exposure of wild type S49 lymphoma cells to somatostatin (which inhibits adenylate cyclase) promotes the translocation of beta AR kinase to an extent comparable to that observed in the presence of the beta agonist isoproterenol or prostaglandin E1. Beta AR kinase activity can be measured using either beta AR or rhodopsin, the retinal receptor for light, as a substrate. The translocation process triggered by somatostatin is rapid, reversible, and is associated with somatostatin receptor desensitization. The latter is apparent as an attenuation of the inhibition by somatostatin of forskolin-stimulated adenylate cyclase activity in membranes of S49 cells preincubated in the presence of the peptide. These results strongly suggest that beta AR kinase is able to phosphorylate and desensitize both stimulatory and inhibitory adenylate cyclase-coupled receptors, thus emerging as a general kinase that regulates the function of different receptors in an agonist-specific fashion.

Animals↗
PubMed1987

The D2-dopamine receptor of anterior pituitary is functionally associated with a pertussis toxin-sensitive guanine nucleotide binding protein.

Dopaminergic inhibition of prolactin release from the anterior pituitary may be mediated through both the adenylate cyclase and Ca2+ mobilization/phosphoinositide pathways. The D2-dopamine receptor of the bovine anterior pituitary has been partially purified by affinity chromatography on CMOS-Sepharose (immobilized carboxymethyleneoximinospiperone). Reinsertion of these partially purified receptor preparations into phospholipid vesicles reconstituted guanine nucleotide-sensitive high affinity agonist binding, agonist-promoted GTPase and 35S-labeled guanosine 5'-O-(thiotriphosphate) [( 35S]GTP gamma S) binding activity in these preparations. Pertussis toxin treatment of the purified receptor preparation abolished agonist-stimulated GTPase and guanine nucleotide-sensitive high affinity agonist binding. These observations suggest that the receptor copurifies with an endogenous, pertussis toxin-sensitive guanine nucleotide binding protein (N). [32P]ADP-ribosylation of affinity-purified D2 receptor preparations by pertussis toxin revealed the presence of a substrate of Mr 39,000-40,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Peptide maps generated using elastase of the [32P]ADP-ribosylated endogenous N protein, transducin, and Ni and No from brain revealed similarities but not identity between the endogenous pituitary N protein and brain Ni and No. Immunoblotting of the partially purified D2 receptor preparations showed an Mr 39,000-40,000 band with an Ni-specific antiserum raised against a synthetic peptide, and with RV3, an No-specific anti-serum, but not with CW6, an antiserum strongly reactive with brain Ni. Several lines of evidence indicate that endogenous pituitary N protein is functionally coupled to the D2 receptor. As measured by [35S]GTP gamma S binding, ratios of 0.2-0.6 mol N protein/mol receptor were observed. Association of N protein with the D2 receptor was increased by agonist pretreatment and decreased by guanine nucleotides. These results suggest that No and/or a form of Ni distinct from the Mr 41,000 pertussis toxin substrate (Ni) is the predominant N protein functionally coupled with the D2-dopamine receptor of anterior pituitary.

Adenylate Cyclase Toxin↗
PubMed1987

Regulation of transmembrane signaling by receptor phosphorylation.

At least two major effects of receptor phosphorylation have been identified--regulation of receptor function, and regulation of receptor distribution. In many cases where phosphorylation directly alters the functions of receptors, this appears to be in a negative direction. Such decreases in receptor activity may reflect reduced ability to interact with biochemical effectors (e.g., the beta-adrenergic receptor, rhodopsin), reduced affinity for binding agonist ligands (EGF,IGF-I, insulin receptors) or reduced enzymatic activity (e.g., tyrosine kinase activity of the insulin or EGF receptor). In all instances, these negative modulations are associated with phosphorylation of serine and/or threonine residues of the receptor proteins. In contrast, the tyrosine kinase receptors also appear to be susceptible to positive modulation by phosphorylation. With these receptors, autophosphorylation of tyrosine residues may lead to enhanced protein-tyrosine kinase activity of the receptors and increased receptor function. In addition, the subcellular distribution of a receptor may be regulated by its phosphorylation status (e.g., the beta-adrenergic receptor, receptors for insulin, EGF, IGF-II, and transferrin). The emerging paradigm is that receptor phosphorylation may in some way promote receptor internalization into sequestered compartments where dephosphorylation occurs. The molecular and cellular mechanisms involved in translating changes in receptor phosphorylation into changes in receptor distribution remain to be elucidated. Moreover, the biological role of receptor internalization may be quite varied. Thus, in the case of the beta-adrenergic receptor, it may serve primarily as a mechanism for bringing the phosphorylated receptors into contact with intracellular phosphatases that dephosphorylate and resensitize it. By contrast, for the transferrin receptor and other receptors involved in receptor-mediated endocytosis, the internalization presumably functions to carry some specific ligand or metabolite into the cell. The role of phosphorylation in regulating receptor function dramatically extends the range of regulatory control of this important covalent modification.

Animals↗
PubMed1987

Functional differences in the beta gamma complexes of transducin and the inhibitory guanine nucleotide regulatory protein.

We have examined the mechanism of inhibition of adenylate cyclase using the purified alpha and beta gamma subunits of bovine brain inhibitory guanine nucleotide regulatory protein (Ni) (i.e., alpha i and beta gamma N) and bovine retinal transducin (alpha T and beta gamma T) in reconstituted phospholipid vesicle systems. The addition of beta gamma N or beta gamma T to lipid vesicles containing the pure stimulatory guanine nucleotide regulatory protein (Ns) from human erythrocytes as well as a resolved preparation of the catalytic moiety (C) of bovine caudate adenylate cyclase results in significant inhibition of guanine nucleotide stimulated cyclase activity (80-90%). The inhibition by these beta gamma subunit complexes appears to fully account for the inhibitory effects observed with holo-Ni or holotransducin. A variety of structure-function comparisons of the beta gamma N and beta gamma T complexes were performed in order to further probe the molecular mechanisms involved in the inhibitory pathway. Whereas the beta subunits of beta gamma N and beta gamma T appear to be very similar, if not identical, on the basis of comparisons of their gel electrophoretic mobility and immunological cross-reactivity, clear differences exist in the apparent structures of gamma N and gamma T. Interestingly, functional differences are observed in the effectiveness of these two beta gamma complexes to inhibit adenylate cyclase activity. Specifically, while both beta gamma N and beta gamma T are capable of effecting significant levels of inhibition of the guanine nucleotide stimulated activities, the beta gamma N complex is consistently more potent than beta gamma T in inhibiting these activities.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclase Inhibitors↗
PubMed1987

Regulation of adrenergic receptor function by phosphorylation. II. Effects of agonist occupancy on phosphorylation of alpha 1- and beta 2-adrenergic receptors by protein kinase C and the cyclic AMP-dependent protein kinase.

The present study was undertaken to determine the ability of protein kinase C and protein kinase A to directly phosphorylate the purified alpha 1- and beta 2-adrenergic receptors (AR). Both the catalytic subunit of protein kinase A and the protein kinase C, purified from bovine heart and pig brain, respectively, are able to phosphorylate the purified alpha 1-AR from DDT1 MF-2 smooth muscle cells. Occupancy of the receptor by an alpha 1 agonist, norepinephrine (100 microM), increases the rate of phosphorylation by protein kinase C but not by protein kinase A. The maximum stoichiometry of phosphorylation obtained is not affected by the agonist and reached 3 mol of PO4/mol of receptor for protein kinase C and 1 mol of PO4/mol of receptor for protein kinase A. The phosphopeptide maps of the trypsinized alpha 1-AR phosphorylated by each kinase differ drastically. The beta 2-AR purified from hamster lungs can also be phosphorylated by the two kinases. In contrast to the alpha 1-AR, the occupancy of the beta 2-AR by the agonist isoproterenol (20 microM) increases the rate of phosphorylation of the beta 2-AR by protein kinase A but not by protein kinase C. The maximum amount of phosphate incorporated into the receptor is not affected in either case by the agonist and reaches 1 mol of PO4/mol of receptor with protein kinase A and 0.4 mol of PO4/mol of receptor with protein kinase C. The phosphopeptide maps of the trypsinized receptor phosphorylated by either kinase reveal similar profiles. Thus, both alpha 1-AR and beta 2-AR are substrates for protein kinase A and protein kinase C. Agonist occupancy of the two receptors facilitates their phosphorylation only by the protein kinase coupled to their own signal transduction pathway. These observations suggest that "feedback" and "cross-system" phosphorylation may represent distinct and differently regulated mechanisms of modulation of receptor function.

Animals↗
PubMed1987

Molecular mechanisms of beta-adrenergic receptor desensitization.

Multiple mechanisms seem to be involved in regulating the responsiveness of hormone receptor-coupled adenylate cyclase systems. These mechanisms at least involve the receptors and nucleotide regulatory proteins. With the recent development of methods for purifying the catalytic unit of the enzyme it will be possible to assess whether it is also a locus for such regulatory phenomena. At least two major pathways of receptor regulation have been uncovered. Homologous desensitization (Fig. 9) involves the uncoupling and translocation of the receptors out of their normal plasma membrane environment. This process sequesters the receptors away from their effector, the regulatory and catalytic components of adenylate cyclase. The site of receptor sequestration is unclear and might lie within the plasma membrane or within the cell. The sequestered receptors can recycle to the cell surface or become down-regulated, perhaps being destroyed within the cell. Phosphorylation of the receptors through a beta-adrenergic receptor kinase appears to be associated with homologous desensitization. This phosphorylation event may serve either to uncouple functionally the receptors or to trigger their sequestration from the cell surface or both. In heterologous desensitization (Fig. 10), receptor function is regulated by phosphorylation in the absence of receptor sequestration or down-regulation. This covalent modification serves to functionally uncouple the receptors, that is, to impair their interactions with the guanine nucleotide regulatory proteins. Several protein kinases seem to be capable of promoting phosphorylation of the receptors including the cAMP-dependent kinase and protein kinase C. In addition to the receptor modification, heterologous desensitization seems to be associated with functional modifications (phosphorylation?) at the level of nucleotide regulatory proteins (Ns and Ni), (Fig. 10). Further studies of the mechanisms of desensitization of adenylate cyclase-coupled receptors are thus likely to help elucidate modes of regulation of a wide variety of receptor-coupled functions in diverse types of cells.

Adenylyl Cyclases↗
PubMed1987

Functional desensitization of the isolated beta-adrenergic receptor by the beta-adrenergic receptor kinase: potential role of an analog of the retinal protein arrestin (48-kDa protein).

The beta-adrenergic receptor kinase is an enzyme, possibly analogous to rhodopsin kinase, that multiply phosphorylates the beta-adrenergic receptor only when it is occupied by stimulatory agonists. Since this kinase may play an important role in mediating the process of homologous, or agonist-specific, desensitization, we investigated the functional consequences of receptor phosphorylation by the kinase and possible analogies with the mechanism of action of rhodopsin kinase. Pure hamster lung beta 2-adrenergic receptor, reconstituted in phospholipid vesicles, was assessed for its ability to mediate agonist-promoted stimulation of the GTPase activity of coreconstituted stimulatory guanine nucleotide-binding regulatory protein. When the receptor was phosphorylated by partially (approximately 350-fold) purified preparations of beta-adrenergic receptor kinase, as much as 80% inactivation of its functional activity was observed. However, the use of more highly purified enzyme preparations led to a dramatic decrease in the ability of phosphorylation to inactivate the receptor such that pure enzyme preparations (approximately 20,000-fold purified) caused only minimal (approximately 1off/- 7%) inactivation. Addition of pure retinal arrestin (48-kDa protein or S antigen), which is involved in enhancing the inactivating effect of rhodopsin phosphorylation by rhodopsin kinase, led to partial restoration of the functional effect of beta-adrenergic receptor kinase-promoted phosphorylation (41 +/- 3% inactivation). These results suggest the possibility that a protein analogous to retinal arrestin may exist in other tissues and function in concert with beta-adrenergic receptor kinase to regulate the activity of adenylate cyclase-coupled receptors.

Animals↗
PubMed1987

The mammalian beta-adrenergic receptor: structural and functional characterization of the carbohydrate moiety.

Mammalian beta-adrenergic receptors are glycoproteins consisting of a single polypeptide chain of Mr approximately 64,000. Treatment of purified [125I]-labeled hamster lung beta-adrenergic receptor with alpha-mannosidase reveals two discrete populations of receptor consistent with previous studies using membrane bound photoaffinity-labeled receptor. Treatment of the [125I]-labeled receptor with endoglycosidase F results initially in the formation of a Mr approximately 57,000 peptide which is further converted to Mr approximately 49,000 suggesting that there are two N-linked carbohydrate chains per receptor polypeptide. Exoglycosidase treatments and lectin chromatography of the [125I]-labeled receptor reveals the presence of two complex type carbohydrate chains (approximately 10% of which are fucosylated) on approximately 45% of the receptors. The remaining approximately 55% of the receptors appear to contain a mixture of carbohydrate chains (possibly high mannose, hybrid and complex type chains). Deglycosylation of the receptor by endoglycosidase F does not appear to alter the binding affinity of the receptor for a variety of beta-adrenergic agonists and antagonists. Moreover, the ability of control, alpha-mannosidase sensitive or insensitive (fractionated on immobilized wheat germ agglutinin) and neuraminidase, alpha-mannosidase or endoglycosidase F treated receptors to interact with the stimulatory guanine nucleotide regulatory protein in a reconstituted system were virtually identical. The deglycosylated receptor was also unaltered in its heat lability as well as its susceptibility to a variety of proteases. These findings demonstrate that the carbohydrate portion of the beta-receptor does not contribute to determining either its specificity of ligand binding or coupling to the adenylate cyclase system.

Adrenergic beta-Agonists↗
PubMed1986

Functional reconstitution of the alpha 2-adrenergic receptor with guanine nucleotide regulatory proteins in phospholipid vesicles.

We describe the successful reconstitution of functional interactions between an inhibitory adenylate cyclase-coupled receptor and various nucleotide-binding regulatory proteins in phospholipid vesicles. The receptor is the alpha 2-adrenergic receptor (alpha 2AR) which has been partially purified (approximately 500-5000-fold) from human platelet membranes. The nucleotide-binding regulatory proteins include purified preparations of human erythrocyte Ni and Ns, bovine retinal transducin and the recently discovered bovine brain No. Addition of the physiologic ligand, epinephrine, to vesicles containing the alpha 2AR and Ni results in stimulation of the GTPase activity in Ni. This stimulation of GTPase activity by epinephrine is prevented in the presence of the alpha-adrenergic antagonist, phentolamine, which indicates that a functional reconstitution of the alpha 2AR and Ni has been established. The maximum turnover number for the alpha 2AR-mediated epinephrine-stimulated GTPase activity in Ni is similar to the maximal turnover numbers obtained for the beta-adrenergic receptor-mediated isoproterenol-stimulated GTPase activity in Ns and the rhodopsin-mediated light-stimulated GTPase activity in transducin (0.5-1.5 mol of Pi released per min per mol of nucleotide regulatory protein). Functional similarities between the alpha 2AR and rhodopsin are observed in their interactions with the various nucleotide-binding regulatory proteins. Thus, both of these receptor proteins are capable of promoting the maximal activation of Ni and No while being much less effective in promoting the activation of Ns. However, there are differences between the alpha 2AR and rhodopsin in their interactions with transducin. Specifically, while rhodopsin will maximally activate transducin, the alpha 2AR is much less effective in promoting this activation (i.e. approximately 20% as effective as rhodopsin). Overall, these results suggest the following specificities of interaction: for rhodopsin, transducin approximately equal to Ni approximately equal to No much greater than Ns; while for alpha 2AR, Ni approximately equal to No greater than transducin greater than or equal to Ns.

Adenylyl Cyclases↗
PubMed1986

Beta-agonist- and prostaglandin E1-induced translocation of the beta-adrenergic receptor kinase: evidence that the kinase may act on multiple adenylate cyclase-coupled receptors.

beta-Adrenergic receptor kinase (beta-AR kinase) is a cytosolic enzyme that phosphorylates the beta-adrenergic receptor only when it is occupied by an agonist [Benovic, J. Strasser, R. H., Caron, M. G. & Lefkowitz, R. J. (1986) Proc. Natl. Acad. Sci. USA 83, 2797-2801.] It may be crucially involved in the processes that lead to homologous or agonist-specific desensitization of the receptor. Stimulation of DDT1MF-2 hamster smooth muscle cells or S49 mouse lymphoma cells with a beta-agonist leads to translocation of 80-90% of the beta-AR kinase activity from the cytosol to the plasma membrane. The translocation process is quite rapid, is concurrent with receptor phosphorylation, and precedes receptor desensitization and sequestration. It is also transient, since much of the activity returns to the cytosol as the receptors become sequestered. Stimulation of beta-AR kinase translocation is a receptor-mediated event, since the beta-antagonist propranolol blocks the effect of agonist. In the kin- mutant of the S49 cells (lacks cAMP-dependent protein kinase), prostaglandin E1, which provokes homologous desensitization of its own receptor, is at least as effective as isoproterenol in promoting beta-AR kinase translocation to the plasma membrane. However, in the DDT1MF-2 cells, which contain alpha 1-adrenergic receptors coupled to phosphatidylinositol turnover, the alpha 1-agonist phenylephrine is ineffective. These results suggest that the first step in homologous desensitization of the beta-adrenergic receptor may be an agonist-promoted translocation of beta-AR kinase from cytosol to plasma membrane and that beta-AR kinase may represent a more general adenylate cyclase-coupled receptor kinase that participates in regulating the function of many such receptors.

Alprostadil↗