A beta-adrenergic receptor kinase-like enzyme is involved in olfactory signal termination.

Abstract

We have previously shown that secondmessenger-dependent kinases (cAMP-dependent kinase, protein kinase C) in the olfactory system are essential in terminating second-messenger signaling in response to odorants. We now document that subtype 2 of the ,B-adrenergic receptor kinase (fARK) is also involved in this process. By using subtype-specific antibodies to .ARK-1 and I3ARK-2, we show that flARK-2 is preferentially expressed in the olfactory epithelium in contrast to 'mdings in most other tissues. Heparin, an inhibitor of DARK, as well as anti-flARK-2 antibodies, (i) completely prevents the rapid decline of second-messenger signals (desensitization) that follows odorant stimulation and (ii) strongly inhibits odorant-induced phosphorylation of olfactory ciliary proteins. In contrast, (6ARK-1 antibodies are without effect. Inhibitors ofprotein kinase A and protein kinase C also block odorant-induced desensitization and phosphorylation. These data suggest that a sequential interplay of secondmessenger-dependent and receptor-specific kinases is functionally involved in olfactory desensitization.The olfactory system responds precisely to iterative stimulation (1) and thus can continuously monitor changes in the odorous environment occurring between consecutive sniffs. This sensory behavior is due to characteristic phasic responses of receptor cells to short odor pulses (2), thereby preventing a brief stimulus from being perceived as continuous smell. The basis for this characteristic feature of olfactory neurons is a rapid termination of the odor-induced primary reaction-i.e., the second-messenger cascade initiated by an odorant-occupied receptor is rapidly turned off (3,4). Recent studies have indicated that olfactory signaling is terminated by uncoupling the transduction cascade; this is accomplished via a negative-feedback reaction controlled by the second messenger elicited upon odor stimulation. Second-messenger-activated protein kinases [protein kinase A (PKA) or protein kinase C (PKC)] presumably phosphorylate elements of the transduction apparatus and thereby uncouple the signaling cascade (5). Subsequent experiments have shown that stimulation with odorants indeed caused significant phosphorylation of olfactory ciliary proteins, and there is some circumstantial evidence indicating that receptor proteins for odorants may be modified via second-messengercontrolled protein kinases (6). The emerging picture for signal termination in olfaction is, in many respects, reminiscent of desensitization phenomena in other systems-notably the visual and hormonal signaling pathways, where phosphorylation of receptors in an active state uncouples the reaction cascade by preventing further stimulation ofG proteins (7,8).For the most widely studied model, the p-adrenergic system, two distinct pathways for phosphorylation and desensitization have recently been discovered. Receptor phosphorylation and cascade uncoupling may be mediated either by a cAMP-dependent protein kinase (PKA) or alternatively by a cAMP-independen...

Phosphorylation and Desensitization of Human Endothelin A and B Receptors

“…Moreover, the paradigm for GRK-initiated desensitization of only agonist-occupied receptors (12) is consonant with the homologous ET A -R and ET B -R desensitization observed in a multitude of experimental systems (5)(6)(7)(8)(9)(10)(11). The rapid rate of receptor/effector desensitization observed in some of these systems (4,5,(7)(8)(9)11) is also consistent with a GRK-initiated mechanism (15,16). In this study, the use of dominant-negative GRK2 to inhibit both agonistinduced phosphorylation and desensitization of the ET A -R demonstrates that these events are indeed mediated by a GRK mechanism (18,21,35,36).…”

“…The GRK family comprises six cloned members, with highly homologous catalytic domains flanked by divergent N and C-terminal domains (14). In model receptor systems, GRK-mediated receptor phosphorylation appears to facilitate the binding of an inhibitory arrestin protein to the receptor, resulting in uncoupling of the receptor from its G protein in the process of homologous desensitization (12), within seconds to minutes of receptor activation (15,16). This GRK-initiated process has been shown to attenuate severely (17,18) or even to terminate (19) signaling through a variety of G protein-coupled receptors.…”

G Protein-coupled Receptor Kinase 3 (GRK3) Gene Disruption Leads to Loss of Odorant Receptor Desensitization

“…In most tissues GRKs 2 and 3 are co-localized, with GRK2 usually being the predominant isoform. We have previously shown the involvement of GRK3 in the desensitization of olfactory signal transduction (5). In olfactory epithelium GRK2 is virtually absent and GRK3 is highly expressed.…”

“…In most tissues examined GRK2 is the predominant form. However, in the olfactory epithelium GRK2 is virtually absent, and GRK3, as well as the second messenger-dependent protein kinases PKA and PKC, is thought to be responsible for the desensitization of olfactory receptors (5), which are members of the GPCR superfamily (6).…”

Regulatory mechanisms that modulate signalling by G-protein-coupled receptors

“…In contrast, overexpression of a dominantnegative βARK1 mutant attenuates desensitization of the β # -AR, the δ-opioid receptor and the κ-opioid receptor [79][80][81]. Treatment of permeabilized cells with heparin, which inhibits βARK1, blocks phosphorylation and desensitization of the β # -AR [82], and treatment of permeabilized olfactory cells with antibodies to βARK1 reduces phosphorylation and desensitization of olfactory receptors [83,84].…”