Comparative rates of desensitization of beta-adrenergic receptors by the beta-adrenergic receptor kinase and the cyclic AMP-dependent protein kinase.

Abstract

ABSTRACTkinases can mediate about the same maximal extent of desensitization. When the desensitization is measured as the loss of signal-transduction capacity of the receptor system (9), the action ofeach ofthe two kinases individually can result in about 50o desensitization. However, PKA-mediated desensitization occurs at about 100-fold lower agonist concentrations than p8ARK-mediated desensitization. Similar conclusions regarding the high agonist sensitivity of PKA-mediated desensitization have been drawn from studies using S49 mouse lymphoma cells defective in PKA (10).In situations where high agonist concentrations are present, such as within the synaptic cleft, the sensitivity of the desensitization response to low agonist concentrations would seem to be of little importance. Instead, the speed at which desensitization is achieved would appear to be more critical. Thus, the most rapid mechanism will be the most important in these systems. The present study describes a kinetic analysis of the three rapid mechanisms leading to ,8-receptor desensitization. Since thus far the critical amino acids of the receptor required for sequestration have not been identified and since, therefore, receptor sequestration cannot be abolished by the mutagenesis approach, we used the technique of inhibition of the three mechanisms by inhibitors (8). The inhibitors used were heparin for BARK (8, 11), the PKA inhibitor peptide (PKI) of the rabbit muscle protein inhibitor for PKA (12), and concanavalin A (Con A) pretreatment of cells for sequestration (13). Although these inhibitors are clearly not entirely specific, we have shown previously (8) that they are specific in the context of P-receptor desensitization-i.e., all of them affect only one of the three rapid desensitization mechanisms. MATERIALS AND METHODSPermeabilization of A431 Cells. Human epidermoid carcinoma A431 cells were grown to -95% confluency in Dulbecco's modified Eagle's medium supplemented with 10%Abbreviations: BARK, B-adrenergic receptor kinase; PKA, protein kinase A; PKI, PKA inhibitor peptide. 6201The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

β‐Agonist‐induced constitutive β2‐adrenergic receptor activity in bovine tracheal smooth muscle

“…Interestingly, the formation of constitutive receptor activity by fenoterol was not only concentration-dependent but also a rapid process; after a 5 min treatment period, a marked depression of the KCl contraction curve already was found, whereas the e ect of 30 min treatment was not further increased by prolonging the incubation period to 18 h. Since activation of bARK and subsequent b-AR phosphorylation are also known to occur rapidly (Roth et al, 1991), the ®ndings would suggest that the fenoterol-induced constitutive activation of the receptor may involve receptor phosphorylation. This proposal would be in line with recent observations by Pei et al (1994) with constitutively active mutant b 2 -ARs, reconstituted in phospholipid vesicles, showing that this receptor is phosphorylated by bARK to a similar extent as the agonist-stimulated wild type receptor.…”

Mechanisms of desensitization of the adrenocorticotropin response to arginine vasopressin in ovine anterior pituitary cells

“…In contrast, changes in AVP concentration in the peripheral circulation are slow and of low magnitude; for example, after hypertonic saline infusion into ewes, there is a steady increase in plasma AVP concentration from 2 to 7 pM over a period of 90 min (Keller-Wood 1994). In the -adrenergic receptor system, GRK-mediated desensitization is very rapid and requires relatively high agonist concentrations, suggesting that this process might be particularly important in environments such as the synaptic cleft, where agonist concentrations increase rapidly to high concentrations (Roth et al 1991). Correspondingly, the PKA-mediated desensitization of the -adrenergic receptor is slower and occurs at lower agonist concentrations, suggesting that it is better suited to the regulation of responsiveness at nonsynaptic receptors ( Roth et al 1991).…”

“…In the -adrenergic receptor system, GRK-mediated desensitization is very rapid and requires relatively high agonist concentrations, suggesting that this process might be particularly important in environments such as the synaptic cleft, where agonist concentrations increase rapidly to high concentrations (Roth et al 1991). Correspondingly, the PKA-mediated desensitization of the -adrenergic receptor is slower and occurs at lower agonist concentrations, suggesting that it is better suited to the regulation of responsiveness at nonsynaptic receptors ( Roth et al 1991). Similarly, differential desensitization of the V1a and V1b receptors may allow for appropriate regulation of the responsiveness to AVP in different tissues where different concentrations and patterns of AVP exposure occur.…”

Rapid desensitization of neonatal rat liver beta-adrenergic receptors. A role for beta-adrenergic receptor kinase.

“…Several studies (15)(16)(17)(18)(19), have suggested that BARK-mediated regulatory mechanisms would be specially important in the presence of high levels of catecholamines, such as at neural synapses. On the contrary, PKA-mediated modulation, which has been shown to be a slower process ( 18), would predominate at lower agonist concentrations; i.e., in response to normal levels ofcirculating catecholamines.…”