Characterization of the cellular trafficking of the apelin receptor and the discovery of a novel apelin antagonist.
Published: 2006
Language: eng
Number of pages: 153
ISBN-13: 9780494219997
Description: The apelin receptor is a member of the rhodopsin family of G protein-coupled receptors, the largest known group of cell-surface mediators of signal transduction. The aim of these studies was to examine the cellular trafficking of the apelin receptor and to generate novel apelin ligands for potential drug discovery and use in characterizing the apelinergic system. A novel nuclear localization was observed for the apelin receptor in human brain. While constitutive nuclear localization was observed for the closely related angiotensin AT1 and bradykinin B2 receptors in HEK-239T cells, the apelin receptor was localized to cell nuclei only in brain tissue and neuronal cell lines. A nuclear localization signal motif in the third intracellular loop of the apelin receptor was identified by point mutagenesis. For the apelin peptide, I hypothesized that a substitution of the carboxyl-terminal phenylalanine was crucial to apelin agonist activity. Apelin-13 analogues were generated and tested for their role on blood pressure regulation. Injections of apelin-13 into spontaneously hypertensive rats lowered mean arterial blood pressure by &sim;60%. Injections of apelin-13(F13A) did not affect blood pressure, though concomitant administration of apelin-13(F13A) blocked hypotensive effects of apelin-13, which revealed apelin-13(F13A) behaved as an apelin-specific antagonist. Finally, I investigated the mechanisms underlying the divergent intracellular trafficking pathways of the apelin receptor after treatment with apelin-13 and apelin-36. My findings revealed beta-arrestin1 dissociated from the apelin-13-internalized receptor while the apelin-36-internalized receptor was trafficked with beta-arrestin1 to intracellular compartments. Apelin-13-internalized receptors recycled to the cell surface within one hour through a Rab4-dependent process while apelin-36-internalized receptors remained sequestered in Rab5-positive endosomes even four hours after agonist removal. Co-expression of Rab7 constitutively targeted the apelin receptor to the lysosomal pathway while Rab11 was not observed to co-localize with the apelin receptor, suggesting that Rab11-mediated slow recycling of the apelin receptor does not occur. Overall, the discovery of a nuclear localization of the apelin receptor and differential agonist-mediated receptor trafficking suggests unanticipated roles for the apelinergic system in cell signalling and function while the generation of an apelin antagonist has produced a candidate for novel drug discovery.
Notes: Source: Dissertation Abstracts International, Volume: 68-01, Section: B, page: 0231.

Thesis (Ph.D.)--University of Toronto, 2006.

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