Gα16/z Chimeras Efficiently Link a Wide Range of G Protein–Coupled Receptors to Calcium Mobilization

Abstract

G protein—coupled receptors (GPCRs) represent a class of important therapeutic targets for drug discovery. The integration of GPCRs into contemporary high-throughput functional assays is critically dependent on the presence of appropriate G proteins. Given that different GPCRs can discriminate against distinct G proteins, a universal G protein adapter is extremely desirable. In this report, the authors evaluated two highly promiscuous Gα16/zchimeras, 16z25 and 16z44, for their ability to translate GPCR activation into Ca2+mobilization using the fluorescence imaging plate reader (FLIPR) and aequorin. A panel of 24 Gs- or Gi-coupled receptors was examined for their functional association with the Gα16/zchimeras. Although most of the GPCRs tested were incapable of inducing Ca2+mobilization upon their activation by specific agonists, the introduction of 16z25 or 16z44 allowed all of these GPCRs to mediate agonist-induced Ca2+mobilization. In contrast, only 16 of the GPCRs tested were capable of using Gα16to mobilize intracellular Ca2+. Analysis of dose-response curves obtained with the δ-opioid, dopamine D1, and Xenopus melatonin Mel1c receptors revealed that the Gα16/zchimeras possess better sensitivity than Gα16in both the FLIPR and aequorin assays. Collectively, these studies help to validate the promiscuity of the Gα16/zchimeras as well as their application in contemporary drug-screening assays that are based on ligand-induced Ca2+mobilization. ( Journal of Biomolecular Screening 2003:39-49)

Differential chemokine activation of CC chemokine receptor 1‐regulated pathways: ligand selective activation of Gα 14‐coupled pathways

“…1). We have previously shown that the 16z44 chimera [17] exhibits robust coupling to numerous G i -coupled receptors [18]. Upon co-expression of CCR1 and 16z44, all chemokines except MIP-1ˇstimulated IP accumulation (Fig.…”

Rapid Uptake and Degradation of CXCL12 Depend on CXCR7 Carboxyl-terminal Serine/Threonine Residues

“…Because chemokine receptors including CXCR4 signal through the G i/o subfamily (28,29), we also used chimeras that were engineered to link G i/o -coupled receptors to Ca 2ϩ mobilization: chimera qi5 consisting of murine G␣ q with the 5 C-terminal amino acids changed from G␣ q to G␣ i2 (30) and chimera 16z44 consisting of human G␣ 16 and rat G␣ z (31,32).…”

“…G protein signaling of the receptor chimeras was then examined by measuring [Ca 2ϩ ] i transients, which is an established tool to study activation of G proteins of the G q/11 subfamily or of chimeric G i/o -G q/11 G proteins (30,32). Stimulation of naive HEK293 cells with CXCL12 produced a subtle AMD3100-sensitive Ca 2ϩ signal (not shown), indicating that the cells expressed functional endogenous CXCR4 receptors.…”

Signal Transducer and Activator of Transcription 3 Activation by the δ-Opioid Receptor via Gα14Involves Multiple Intermediates

“…1A 16 . The identity of the PTX-insensitive G protein is unlikely to be G q because DOR is incapable of interacting with G q (Liu et al, 2003).…”