Five-transmembrane domains appear sufficient for a G protein-coupled receptor: Functional five-transmembrane domain chemokine receptors (signal transductionydesensitizationyinternalization)

The putative seven-transmembrane (TM) domains have been the structural hallmark for the superfam- ily of heterotrimeric G protein-coupled receptors (GPCRs) that regulate a variety of cellular functions by mediating a large number of extracellular signals. Five-TM GPCR mu- tants of chemokine receptor CCR5 and CXCR4, the N- terminal segment of which connected directly to TM3 as a result of a deletion of TM1-2 and the first intracellular and extracellular loops, have been obtained in this study. Laser confocal microscopy and f low cytometry analysis revealed that these five-TM mutant GPCRs were expressed stably on the cell surface after transfection into human embryonic kidney 293 cells. The five-TM CCR5 and CXCR4 functioned as normal chemokine receptors in mediating chemokine- stimulated chemotaxis, Ca 21 inf lux, and activation of pertus- sis toxin-sensitive G proteins. Like the wild-type GPCRs, the five-TM mutant receptors also underwent agonist-dependent internalization and desensitization and were subjected to regulation by GPCR kinases and arrestins. Our study indi- cates that five-TM domains, at least in the case of CCR5 and CXCR4, appear to meet the minimum structural requirements for a functional GPCR and suggests possible existence of functional five-TM GPCRs in nature during evolution.