Work

Gαs directly drives PDZ-RhoGEF signaling to Cdc42

Year: 2020

Type: article

Abstract: Gα proteins promote dynamic adjustments of cell shape directed by actin-cytoskeleton reorganization via their respective RhoGEF effectors. For example, Gα 13 binding to the RGS-homology (RH) domains of several RH-RhoGEFs allosterically activates these proteins, causing them to expose their catalytic Dbl-homology (DH)/pleckstrin-homology (PH) regions, which triggers downstream signals. However, whether additional Gα proteins might directly regulate the RH-RhoGEFs was not known. To explore this question, we first examined the morphological effects of expressing shortened RH-RhoGEF DH/PH constructs of p115RhoGEF/ARHGEF1, PDZ-RhoGEF (PRG)/ARHGEF11, and LARG/ARHGEF12. As expected, the three constructs promoted cell contraction and activated RhoA, known to be downstream of Gα 13 . Intriguingly, PRG DH/PH also induced filopodia-like cell protrusions and activated Cdc42. This pathway was stimulated by constitutively active Gα s (Gα s Q227L), which enabled endogenous PRG to gain affinity for Cdc42. A chemogenetic approach revealed that signaling by G s -coupled receptors, but not by those coupled to G i or G q , enabled PRG to bind Cdc42. This receptor-dependent effect, as well as CREB phosphorylation, was blocked by a construct derived from the PRG:Gα s -binding region, PRG-linker. Active Gα s interacted with isolated PRG DH and PH domains and their linker. In addition, this construct interfered with Gα s Q227L's ability to guide PRG's interaction with Cdc42. Endogenous G s -coupled prostaglandin receptors stimulated PRG binding to membrane fractions and activated signaling to PKA, and this canonical endogenous pathway was attenuated by PRG-linker. Altogether, our results demonstrate that active Gα s can recognize PRG as a novel effector directing its DH/PH catalytic module to gain affinity for Cdc42.

Source: Journal of Biological Chemistry

DOI: 10.1074/jbc.ac120.015204