Protein kinase C potentiates isoproterenol‐mediated cyclic AMP production without modifying the homologous desensitization process in J774 cells

Abstract

The J774 murine macrophage cells possess a P2-adrenergic receptor coupled to adenylate cyclase, which can be regulated by homologous desensitization. Stimulation of protein kinase C by phorbol esters or oleoyl acetyl glycerol potentiates two-to-threefold the isoproterenol-induced cyclic AMP accumulation. These promoters act at a post-receptor level, since the number and affinity of the P-adrenergic receptors, measured by use of the hydrophilic ligand [3H]CGP-12 177, are not modified. In addition, the effect of cholera toxin is similarly increased and pretreatment of the cells with pertussis toxin prevents the action of phorbol esters. On the other hand, these promoters are ineffective on isoproterenol-induced desensitization and the rates of receptor segregation and recovery remain unchanged. Therefore, protein kinase C modulates the isoproterenol-stimulated adenylate cyclase, whereas it is inactive on the homologous desensitization process.The phorbol esters are well-known tumor promoters which stimulate the CaZ + and phospholipid-dependent protein kinase C. They substitute for diacylglycerol, which is the natural activator of protein kinase C and is produced from hormone and growth-factor-induced turnover of polyphosphoinositides [l].Recently, it has been demonstrated that the phorbol esters can interfere with the hormone-mediated adenylate cyclase activation. The data concerning p-adrenergic-mediated stimulation depend on the cell type studied. Thus, in some cells the tumor promoters induce P-adrenergic desensitization [2 -161, while in others they potentiate the response to isoproterenol [17-241. In addition, in cell-free systems protein kinase C either increases [20, 251 or decreases [8, 261 the basal and stimulated adenylate cyclase. The molecular mechanisms of these effects are actively investigated. The data demonstrate that the number and affinity of P-adrenergic receptors remain unchanged [5,9,13, 18, 20, 221 has demonstrated that the catalytic unit of adenylate cyclase is phosphorylated in vivo by phorbol esters [29].On the other hand, few studies have investigated the action of protein kinase C on the homologous desensitization of padrenergic receptors [16, 241. Recent work has demonstrated that this is a cyclic-AMP-independent phenomenon, related to the phosphorylation of B-receptors by a specific kinase (review in [30]). The aim of our study was to investigate the effect of phorbol esters on the two types of the action of j-agonists: the stimulation of cyclic AMP production and the homologous desensitization. We took advantage of the availability of the hydrophilic P-adrenergic ligand, [3H]CGP-12 177, which is membrane-impermeant and allows for the investigation of surface receptors on intact cells [31].We used in this study a continuous cell line of murine macrophages, 5774 [32, 331, which is sensitive both to cyclic AMP derivatives [34, 351 and to phorbol esters [36].Our data demonstrate that (a) protein kinase C activation by phorbol esters or oleoyl acetyl glycerol potentiates the isopr...

Biphasic regulation of macrophage attachment by activators of cyclic adenosine monophosphate‐dependent kinase and protein kinase C

“…Preincubation of the cells with OAG resulted in a potentiation of the stimulatory effect of PGEl on cAMP production (Fig. 71, as was described for isoproterenol (Chambaut-Guerin and Thomopoulos, 1987). Under the same experimental conditions, PGEl-induced adhesiveness was clearly inhibited by preincubation with OAG (data not shown).…”

“…71, as was described for isoproterenol (Chambaut-Guerin and Thomopoulos, 1987). Under the same experimental conditions, PGEl-induced adhesiveness was clearly inhibited by preincubation with OAG (data not shown).…”

“…The assays were carried out in triplicate. cAMP accumulation cAMP cell content was measured by radioimmunoassay after trichloroacetic acid extraction (Chambaut-Guerin and Thomopoulos, 1987).…”

Gravin-mediated Formation of Signaling Complexes in β2-Adrenergic Receptor Desensitization and Resensitization

“…Protein kinase C has been shown to modulate the function of several G-protein-linked receptors, and consensus sites for phosphorylation have been identified in many of these receptors. Although a few early studies suggested that activation of protein kinase C desensitizes the ␤ 2 -adrenergic receptors (38,39), most subsequent studies reported either no effect of protein kinase C activation on desensitization (40,41) or frank potentiation of ␤ 2 -adrenergic receptor signaling rather than desensitization (42)(43)(44)(45)(46)(47)(48). Protein kinase C is associated in complex with the receptor-gravin complex in unstimulated cells.…”

Cyclic adenosine monophosphate and diacylglycerol. Mutually inhibitory second messengers in cultured rat inner medullary collecting duct cells.

“…For example, cAMP has been shown to inhibit phosphoinositide hydrolysis in platelets (2) and neutrophils (3), but it has no effect in monocytes (4). Conversely, activation of protein kinase C, one of the major consequences of phosphoinositide hydrolysis, has been reported to inhibit hormone-stimulated AC activity in some tissues (5)(6)(7)(8), while it potentiates AC activity in other tissues (9)(10)(11)(12).…”