Control of feeding behavior in C. elegans by human G protein-coupled receptors permits screening for agonist-expressing bacteria
G protein-coupled receptors (GPCRs) have a key role in many biological processes and are important drug targets for many human diseases. Therefore, understanding the molecular interactions between GPCRs and their ligands would improve drug design. Here, we describe an approach that allows the rapid identification of functional agonists expressed in bacteria. Transgenic Caenorhabditis elegans expressing the human chemokine receptor 5 (CCR5) in nociceptive neurons show avoidance behavior on encounter with the ligand MIP-1α and avoid feeding on Escherichia coli expressing MIP-1α compared with control bacteria. This system allows a simple activity screen, based on the distribution of transgenic worms in a binary food-choice assay, without a requirement for protein purification or tagging. By using this approach, a library of 68 MIP-1α variants was screened, and 13 critical agonist residues involved in CCR5 activation were identified, four of which (T8, A9, N22, and A25) have not been described previously, to our knowledge. Identified residues were subsequently validated in receptor binding assays and by calcium flux assays in mammalian cells. This approach serves not only for structure/function studies as demonstrated, but may be used to facilitate the discovery of agonists within bacterial libraries.
|Keywords||Caenorhabditis elegans, Escherichia coli, G protein coupled receptor, animal, article, avoidance behavior, biosynthesis, calcium transport, chemistry, chemokine receptor CCR5, controlled study, drug antagonism, drug potentiation, feeding behavior, gene library, genetics, human, isolation and purification, ligand, macrophage inflammatory protein 1alpha, metabolism, nerve cell, nonhuman, physiology, priority journal, protein binding, protein conformation, protein purification, screening, transgene, transgenic animal, transgenics|
|Persistent URL||dx.doi.org/10.1073/pnas.0803290105, hdl.handle.net/1765/14554|
|Journal||Proceedings of the National Academy of Sciences of the United States of America|
Teng, M.S, Shadbolt, P, Fraser, A.G, Jansen, G, & McCafferty, J. (2008). Control of feeding behavior in C. elegans by human G protein-coupled receptors permits screening for agonist-expressing bacteria. Proceedings of the National Academy of Sciences of the United States of America, 105(39), 14826–14831. doi:10.1073/pnas.0803290105