Characterization of SEQ-9 an Orphan G protein-Coupled Receptor
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Authors
Mandrekar, Shweta
Issue Date
2001
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
Current research in pharmaceuticals is primarily focused on cell signaling pathways.
G-proteins (GTP binding proteins) are a particularly favorite drug target because of their
ubiquitous expression and the vast number of agonists that bind to them. Due to these factors they are implicated in a whole array of physiological diseases and disorders. SEQ-9, a novel G-protein coupled receptor (GPCR), was identified from a proprietary human genomic database and is characterized by common motifs that are shared among GPCRs. The goal of the project was to clone a novel full-length GPCR ID'd from a proprietary database. The function of the resulting construct was characterized by assays commonly used to study GPCR function in cell signaling pathways. A BLAST search indicated that SEQ-9 was the human homolog of the rat EDG-8, sharing a 87.7% identity at the amino acid level. Edg-8 has been shown to have SIP (Spingosine-I-Phosphate) as its ligand therefore, this information raised the possibility of SIP as a ligand for SEQ-9. An adenylyl cyclase assay showed that SIP induced an inhibition of forskolin-stimulated intracellular cyclic AMP levels suggesting coupling through a G.-coupled signaling pathway. A FLIPR () assay was also performed to monitor levels of calcium within the cells. A four to five-fold increase in calcium mobilization was seen when the cells were stimulated with SIP, suggesting coupling through a Gq signaling pathway. These experiments strongly suggest that SIP is a ligand for SEQ-9, and that this GPCR couples through both the Gi and Gq signaling pathways.
Description
iv, 37 p.
Citation
Publisher
Kalamazoo College
License
U.S. copyright laws protect this material. Commercial use or distribution of this material is not permitted without prior written permission of the copyright holder.