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Differential targeting of heterotrimeric G protein versus ?-arrestin signaling are emerging concepts in G protein-coupled receptor (GPCR) research and drug discovery, and biased engagement by GPCR ligands of either ?-arrestin or G protein pathways has been disclosed. Herein we report on a new mechanism of ligand bias to titrate the signaling specificity of a cell-surface GPCR. Using a combination of biomolecular and virtual screening, we identified the small-molecule modulator Gue1654, which inhibits G?? but not G? signaling triggered upon activation of G?i-?? by the chemoattractant receptor OXE-R in both recombinant and human primary cells. Gue1654 does not interfere nonspecifically with signaling directly at or downstream of G??. This hitherto unappreciated mechanism of ligand bias at a GPCR highlights both a new paradigm for functional selectivity and a potentially new strategy to develop pathway-specific therapeutics. [ABSTRACT FROM AUTHOR]

Label-free biosensor technology based on dynamic mass redistribution (DMR) of cellular constituents promises to translate GPCR signaling into complex optical 'fingerprints' in real time in living cells. Here we present a strategy to map cellular mechanisms that define label-free responses, and we compare DMR technology with traditional second-messenger assays that are currently the state of the art in GPCR drug discovery. The holistic nature of DMR measurements enabled us to (i) probe GPCR functionality along all four G-protein signaling pathways, something presently beyond reach of most other assay platforms; (ii) dissect complex GPCR signaling patterns even in primary human cells with unprecedented accuracy; (iii) define heterotrimeric G proteins as triggers for the complex optical fingerprints; and (iv) disclose previously undetected features of GPCR behavior. Our results suggest that DMR technology will have a substantial impact on systems biology and systems pharmacology as well as for the discovery of drugs with novel mechanisms. [ABSTRACT FROM AUTHOR]

In spite of the crucial role of heterotrimeric G proteins as molecular switches transmitting signals from G protein-coupled receptors, their selective manipulation with small molecule, cell-permeable inhibitors still remains an unmet challenge. Here, we report that the small molecule BIM-46187, previously classified as pan-G protein inhibitor, preferentially silences Gαq signaling in a cellular context-dependent manner. Investigations into its mode of action reveal that BIM traps Gαq in the empty pocket conformation by permitting GDP exit but interdicting GTP entry, a molecular mechanism not yet assigned to any other small molecule Gα inhibitor to date. Our data show that Gα proteins may be "frozen" pharmacologically in an intermediate conformation along their activation pathway and propose a pharmacological strategy to specifically silence Gα subclasses with cell-permeable inhibitors. • The pan-G protein inhibitor BIM-46187 is identified as Gαq preferring • BIM-46187 traps Gαq proteins in the empty pocket conformation • BIM-46187 uncouples high-affinity agonist binding from agonist function • Research tools assess G-protein-mediated cell responses Schmitz et al. show that BIM-46187, previously classified as a pan G protein inhibitor, may instead serve to specifically silence Gαq proteins by "freezing" Gαq in its empty pocket conformation, a conformational intermediate along the Gα activation pathway. [ABSTRACT FROM AUTHOR]