Sphingosine 1-phosphate-mediated α1B-adrenoceptor desensitization and phosphorylation. Direct and paracrine/autocrine actions

Abstract

Sphingosine-1-phosphate-induced α1B-adrenergic receptor desensitization and phosphorylation was studied in rat-1 fibroblasts stably expressing enhanced green fluorescent protein-tagged adrenoceptors. Sphingosine-1-phosphate induced adrenoceptor desensitization and phosphorylation through a signaling cascade that involved phosphoinositide 3-kinase and protein kinase C activities. The autocrine/paracrine role of sphingosine-1-phosphate was also studied. It was observed that activation of receptor tyrosine kinases, such as insulin growth factor-1 (IGF-I) and epidermal growth factor (EGF) receptors increased sphingosine kinase activity. Such activation and consequent production of sphingosine-1-phosphate appears to be functionally relevant in IGF-I- and EGF-induced α1B-adrenoceptor phosphorylation and desensitization as evidenced by the following facts: a) expression of a catalytically inactive (dominant-negative) mutant of sphingosine kinase 1 or b) S1P1 receptor knockdown markedly reduced this growth factor action. This action of sphingosine-1-phosphate involves EGF receptor transactivation. In addition, taking advantage of the presence of the eGFP tag in the receptor construction, we showed that S1P was capable of inducing α1B-adrenergic receptor internalization and that its autocrine/paracrine generation was relevant for internalization induced by IGF-I. Four distinct hormone receptors and two autocrine/paracrine mediators participate in IGF-I receptor- α1B-adrenergic receptor crosstalk.

Protein Kinase C Activation Promotes α1B-Adrenoceptor Internalization and Late Endosome Trafficking through Rab9 Interaction. Role in Heterologous Desensitization

“…PMA and S1P induced a 1B -AR internalization that was markedly diminished by the same treatments, i.e., PKC inhibition, PKC downregulation, and the expression of a GDP-locked dominant-negative Rab9 mutant. These data are consistent with the current idea that PKC is a major mediator of a 1B -AR heterologous desensitization, including receptor phosphorylation and internalization; this can be induced by S1P (Castillo-Badillo et al, 2012 and by a large variety of hormones and neurotransmitters acting through different families of receptors [see Castillo-Badillo et al (2012; García-Sáinz et al (2000; and references therein]. A working model is presented in Fig.…”

“…Fura-2/AM [bis(acetoxymethyl) 2,29-((2-(5-((acetoxymethoxy)carbonyl)oxazol-2-yl)-5-(2-(2-(bis(2-(acetoxymethoxy)-2-oxoethyl)amino)-5-methylphenoxy)ethoxy)benzofuran-6-yl)azanediyl)diacetate]; PubChem 3364574), Dulbecco's modified Eagle's medium, fetal bovine serum albumin, trypsin, antibiotics, and other reagents employed for cell culture were from Life Technologies/ ThermoFisher Scientific. Other reagents were from previously described sources (Castillo-Badillo et al, 2012.…”

“…a 1B -ARs function is tightly regulated and their signaling is markedly attenuated upon sustained agonist activation (homologous desensitization), as well as in response to unrelated agents (heterologous desensitization) acting either through other G protein-coupled receptors, receptor tyrosine kinases, and nuclear receptors, or by direct stimulation of protein kinases such as PKC (García-Sáinz et al, 2000. Agonist-induced a 1B -AR phosphorylation and desensitization is mainly the result of the action of G protein-coupled receptor kinases, whereas second messenger-activated kinases, particularly PKC, are the major mediators of the heterologous process (Diviani et al, 1996(Diviani et al, , 1997García-Sáinz et al, 2000Castillo-Badillo et al, 2012). Receptor phosphorylation appears to be an early step in desensitization, as it triggers recruitment of b-arrestins, clathrin, and other proteins, such as Rab GTPases, which participate in receptor internalization (Lefkowitz, 2013).…”

α1B-Adrenergic Receptors Differentially Associate with Rab Proteins during Homologous and Heterologous Desensitization

“…α 1B -Adrenergic receptor desensitization, phosphorylation, and internalization has been extensively studied and major differences have been observed in these processes when they are triggered either by adrenergic agonists (homologous) or by unrelated agents (heterologous) acting through other GPCRs, receptor tyrosine kinases, nuclear receptors or by direct activation of protein kinase C [ 40 – 57 ]. Among the GPCRs whose activation induce α 1B -adrenergic receptor phosphorylation and internalization is the sphingosine 1-phosphate S1P1 receptor [ 42 ]. Sphingosine 1-phosphate is a bioactive lipid that functions as a paracrine / autocrine mediator [ 58 ], and through the S1P1 receptor plays cardinal roles in the intense crosstalk that takes place between receptor tyrosine kinases and GPCRs [ 42 , 43 , 58 ].…”

Sphingosine-1-phosphate signalling as a therapeutic target for patients with abnormal glucose metabolism and ischaemic heart disease

“…Similarly, it has been reported that cAMP is involved in desensitization of the b 1 -adrenergic receptor as well as the b 2 -adrenergic receptor, probably via direct phosphorylation of the receptors by the cAMP-dependent protein kinase. 48,49 Castillo-Badillo et al 50 showed that activation of S1P 1 receptors may induce a 1B -AR phosphorylation and desensitization via a mechanism that may involve phosphoinositide 3-kinase (PI3K) and PKC signalling. In the cardiovascular system, these observations suggest that activation of S1P signalling may represent an additional potential protective mechanism as it is known that a 1B -ARs may regulate several processes, such as cardiac muscle and arteriolar smooth muscle contraction, and be implicated in other pathological processes, such as cardiac hypertrophy or ischaemia-induced cardiac arrhythmias and the genesis/ maintenance of hypertension.…”