Deletion of the distal COOH-terminus of the A_2B adenosine receptor switches agonist-induced internalisation to an arrestin- and clathrin-independent pathway and inhibits receptor recycling

Mundell, S.J., Matharu, A.-L., Palmer, T.M., Benovic, J.L. and Kelly, E. (2010) Deletion of the distal COOH-terminus of the A_2B adenosine receptor switches agonist-induced internalisation to an arrestin- and clathrin-independent pathway and inhibits receptor recycling. British Journal of Pharmacology, 159(3), pp. 518-533. (doi: 10.1111/j.1476-5381.2009.00598.x)

Abstract

<b>Background and purpose:</b>  We have investigated the effect of deletions of a postsynaptic density, disc large and zo-1 protein (PDZ) motif at the end of the COOH-terminus of the rat A_2B adenosine receptor on intracellular trafficking following long-term exposure to the agonist 5′-(N-ethylcarboxamido)-adenosine.<p></p> <b>Experimental approach:</b>  The trafficking of the wild type A_2B adenosine receptor and deletion mutants expressed in Chinese hamster ovary cells was studied using an enzyme-linked immunosorbent assay in combination with immunofluorescence microscopy.<p></p> <b>Key results:</b>  The wild type A_2B adenosine receptor and deletion mutants were all extensively internalized following prolonged treatment with NECA. The intracellular compartment through which the Gln³²⁵-stop receptor mutant, which lacks the Type II PDZ motif found in the wild type receptor initially trafficked was not the same as the wild type receptor. Expression of dominant negative mutants of arrestin-2, dynamin or Eps-15 inhibited internalization of wild type and Leu³³⁰-stop receptors, whereas only dominant negative mutant dynamin inhibited agonist-induced internalization of Gln³²⁵-stop, Ser³²⁶-stop and Phe³²⁸-stop receptors. Following internalization, the wild type A_2B adenosine receptor recycled rapidly to the cell surface, whereas the Gln³²⁵-stop receptor did not recycle.<p></p> <b>Conclusions and implications:</b>  Deletion of the COOH-terminus of the A2B adenosine receptor beyond Leu³³⁰ switches internalization from an arrestin- and clathrin-dependent pathway to one that is dynamin dependent but arrestin and clathrin independent. The presence of a Type II PDZ motif appears to be essential for arrestin- and clathrin-dependent internalization, as well as recycling of the A_2B adenosine receptor following prolonged agonist addition.<p></p>

• Item Type: Articles
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Palmer, Dr Timothy
• Authors: Mundell, S.J., Matharu, A.-L., Palmer, T.M., Benovic, J.L., and Kelly, E.
• College/School: College of Medical Veterinary and Life Sciences; College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health
• Journal Name: British Journal of Pharmacology
• Journal Abbr.: Br. J. Pharmacol.
• Publisher: Wiley
• ISSN: 0007-1188
• ISSN (Online): 1476-5381
• Published Online: 28 January 2010

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Project Code	Award No	Project Name	Principal Investigator	Funder's Name	Funder Ref	Lead Dept
49858	1	EPAC1 and ERK-dependent activation of C/EBP transcription factors: a new cyclic AMP-activated anti-inflammatory gene expression module in vascular endothelial cells	Timothy Palmer	British Heart Foundation (BHF)	PG/08/125/26415	RI CARDIOVASCULAR & MEDICAL SCIENCES