Potent and Selective Peptide-based Inhibition of the G Protein Gαq

• Published in: The Journal of biological chemistry 2016-12, Vol.291 (49), p.25608-25616
• Main Authors: Charpentier, Thomas H., Waldo, Gary L., Lowery-Gionta, Emily G., Krajewski, Krzysztof, Strahl, Brian D., Kash, Thomas L., Harden, T. Kendall, Sondek, John
• Format: Article
• Language: English
• Subjects: Animals; Cell-Penetrating Peptides - chemistry; Cell-Penetrating Peptides - pharmacology; G protein; GTP-Binding Protein alpha Subunits, Gq-G11 - antagonists & inhibitors; GTP-Binding Protein alpha Subunits, Gq-G11 - genetics; GTP-Binding Protein alpha Subunits, Gq-G11 - metabolism; GTP-Binding Protein beta Subunits - genetics; GTP-Binding Protein beta Subunits - metabolism; GTP-Binding Protein gamma Subunits - genetics; GTP-Binding Protein gamma Subunits - metabolism; HEK293 Cells; Humans; inhibitor; Methods and Resources; Mice; p63; peptide interaction; phospholipase C; Phospholipase C beta - genetics; Phospholipase C beta - metabolism; Prefrontal Cortex - metabolism; Protein Structure, Secondary
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M116.740407

In contrast to G protein-coupled receptors, for which chemical and peptidic inhibitors have been extensively explored, few compounds are available that directly modulate heterotrimeric G proteins. Active Gαq binds its two major classes of effectors, the phospholipase C (PLC)-β isozymes and Rho guanine nucleotide exchange factors (RhoGEFs) related to Trio, in a strikingly similar fashion: a continuous helix-turn-helix of the effectors engages Gαq within its canonical binding site consisting of a groove formed between switch II and helix α3. This information was exploited to synthesize peptides that bound active Gαqin vitro with affinities similar to full-length effectors and directly competed with effectors for engagement of Gαq. A representative peptide was specific for active Gαq because it did not bind inactive Gαq or other classes of active Gα subunits and did not inhibit the activation of PLC-β3 by Gβ1γ2. In contrast, the peptide robustly prevented activation of PLC-β3 or p63RhoGEF by Gαq; it also prevented G protein-coupled receptor-promoted neuronal depolarization downstream of Gαq in the mouse prefrontal cortex. Moreover, a genetically encoded form of this peptide flanked by fluorescent proteins inhibited Gαq-dependent activation of PLC-β3 at least as effectively as a dominant-negative form of full-length PLC-β3. These attributes suggest that related, cell-penetrating peptides should effectively inhibit active Gαq in cells and that these and genetically encoded sequences may find application as molecular probes, drug leads, and biosensors to monitor the spatiotemporal activation of Gαq in cells.