Use this URL to cite or link to this record in EThOS:
Title: Interaction of differentially-coupled GPCRs resulting in enhanced intracellular Ca2+ signalling
Author: Werry, Tim
Awarding Body: University of Leicester
Current Institution: University of Leicester
Date of Award: 2002
Availability of Full Text:
Access from EThOS:
Access from Institution:
The superfamily of G-protein-coupled receptors (GPCRs) represents a major group of cellular receptors that convert extracellular signals from hormones, neurotransmitters and pharmacological ligands into cellular responses. These receptors instigate intracellular signalling by activating heterotrimeric GTP-binding proteins (G-proteins), which subsequently regulate effectors including ion channels and enzymes involved in second messenger production. Examples of the activation of one type of GPCR influencing the activity of a different type of GPCR are becoming increasingly frequent. Such interactions have far-reaching implications, potentially having significance in diverse patho-physiological circumstances. Many chemokines signal via GPCRs, including interleukin-8 and GROcc acting through the Gocj-coupled CXC chemokine receptor 2 (CXCR2). This receptor stimulates pertussis-toxin sensitive Ca2+ responses (i.e. via Ga{) in many native cell types, but not when expressed recombinantly in an immortalized cell line. This study shows that stimulation of endogenous Gaq-coupled P2Y nucleotide receptors in CXCR2-transfected human embryonic kidney (HEK) cells allows a subsequent robust Ca2+ response to stimulation of CXCR2. This interaction required continued nucleotide receptor activation and was dependent on the activation of Gcti G-proteins. Extracellular Ca was not required, indicating release of additional Ca2+ from internal stores following CXCR2 stimulation. However, the extent of potentiation induced by P2Y2 receptors was markedly different to that induced by P2Y1 receptors, a difference attributable to the rapid and full desensitization of P2Y1 receptors. Co- stimulation of CXCR2 and P2Y2 receptors also potentiated phosphoinositide generation, suggesting an involvement of phospholipase C (PLC) and InsP3 in the crosstalk. A similar crosstalk phenomenon occurs between Gaq-coupled M3 muscarinic- and Gocs-coupled (32 adrenoceptors. However, the muscarinic M3 receptor-(32-adrenoceptor interaction is not observed at the phosphoinositide level, indicating two divergent mechanisms, although it was shown that protein kinase A activation was not required. The involvement of the PLCp/InsP3 signalling pathway was supported by the inhibitory effects of the putative InsP3 receptor blocker, 2-aminoethoxy-diphenylborane. That additional Ca2+ stores could be accessed simply by increasing InsP3 production was confirmed by the demonstration of quantal Ca2+ release. The possibility that potentiation was due to co-operativity between Gocq and G(3y acting on PLCP was supported by the demonstration that inhibition of Gccq by regulator of G-protein signalling 2 (RGS2), or the sequestration of Gpy by Gat, markedly attenuated the response to CXCR2 stimulation. In addition, the physiological relevance of these studies was examined in a variety of primary cells expressing both P2Y receptors and CXCR2 endogenously. Similar crosstalk was not observed in any of these cell types, and the potential reasons for this apparent discrepancy between the recombinant system and the native system is discussed. However, this work represents a significant advance in the understanding of a potential mechanism of interaction by which GPCRs can modulate the intracellular Ca2+ signalling of another receptor.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available