Chemokines are detected by cell surface receptors that are members of the G protein coupled receptor (GPCR) family. GPCR proteins serve a wide range of functions, making them attractive targets for therapeutic intervention. Most commonly, the mechanism of ligand binding and receptor activation is studied in model cellular systems through transient expression of the proteins. However, an increasingly large number of mammalian GPCRs are being investigated using yeast as an alternative system. Yeast is an attractive expression host for studying mammalian GPCRs because the signaling pathway is conserved in both organisms, which allows heterologously expressed mammalian GPCR proteins to be coupled to the endogenous pheromone pathway. Yeast offers other advantages over cellular systems, including a faster growth rate, lower experimental cost, and ease of handling. The molecular biology tools are also well developed for yeast, which further facilitates the development of highly sensitive reporter assays in yeast that may be used to measure GPCR activity in high throughput.

In this study, we report the construction of a yeast-based screening system for the neutrophil chemokine receptors. Neutrophils are recruited to a site of injury by the various chemokines secreted by the tissue cells and other leukocytes, yet excessive activation of neutrophil causes a variety of inflammatory disorders, including Crohn's disease and psoriasis. Thus, an efficient screening system to identify novel inhibitors of neutrophil activation would be therapeutically useful. We have expressed the human neutrophil chemokine receptor, CXCR1, in engineered yeast. In our system, the activation of CXCR1 is coupled to the synthesis of various reporter gene products, including LacZ and URA3. This system forms the foundation of high throughput screens for small molecule CXCR1 agonists and antagonists using high density plates. One potential readout system includes measurement of OD600 to monitor cell survival, which allows easy identification of active compounds. Since the paradigm of the assay is invariant with respect to the exact identity of the receptor, similar assays may be developed to find active compounds against other therapeutic GPCRs, as well as to study receptor-ligand interaction in high throughput.