Structure of the Regulator of G Protein Signaling 8 (RGS8)-Gαq Complex

Abstract

Regulator of G protein signaling (RGS) proteins interact with activated G␣ subunits via their RGS domains and accelerate the hydrolysis of GTP. Although the R4 subfamily of RGS proteins generally accepts both G␣ i/o and G␣ q/11 subunits as substrates, the R7 and R12 subfamilies select against G␣ q/11 . In contrast, only one RGS protein, RGS2, is known to be selective for G␣ q/11 . The molecular basis for this selectivity is not clear. Previously, the crystal structure of RGS2 in complex with G␣ q revealed a non-canonical interaction that could be due to interfacial differences imposed by RGS2, the G␣ subunit, or both. To resolve this ambiguity, the 2.6 Å crystal structure of RGS8, an R4 subfamily member, was determined in complex with G␣ q . RGS8 adopts the same pose on G␣ q as it does when bound to G␣ i3 , indicating that the non-canonical interaction of RGS2 with G␣ q is due to unique features of RGS2. Based on the RGS8-G␣ q structure, residues in RGS8 that contact a unique ␣-helical domain loop of G␣ q were converted to those typically found in R12 subfamily members, and the reverse substitutions were introduced into RGS10, an R12 subfamily member. Although these substitutions perturbed their ability to stimulate GTP hydrolysis, they did not reverse selectivity. Instead, selectivity for G␣ q seems more likely determined by whether strong contacts can be maintained between ␣6 of the RGS domain and Switch III of G␣ q , regions of high sequence and conformational diversity in both protein families.Many transmembrane signaling events are transduced inside the cell by heterotrimeric G proteins, which are activated by cell surface G protein-coupled receptors. G protein-coupled receptors stimulate the exchange of bound GDP for GTP on the G␣ subunit, which then separates from the G␤␥ subunits and interacts with downstream effectors (1). After hydrolyzing GTP to GDP, the G␣ subunit is deactivated and is rapidly sequestered by G␤␥. In biological processes, such as the visual response, deactivation of G␣ has been observed at much faster rates than those measured for isolated G␣ subunits in vitro (2, 3). This discrepancy helped lead to the discovery of a family of GTPaseactivating proteins (GAPs), 2 now known as regulator of G protein signaling (RGS) proteins (4 -6). RGS proteins contain a conserved helical domain called the RGS domain that directly binds to the three switch regions (SwI-III) of the G␣ subunit and stabilizes them in a transition state conformation (7).RGS domains are divided into four subfamilies based on sequence homology and substrate preference: RZ, R4, R7, and R12 (8). All utilize G␣ i/o subunits as substrates, although some RZ members seem selective for G␣ z subunits (9). A recent study using surface plasmon resonance indicated that the RGS domains that belong to the R7 and R12 subfamilies bind weakly or not at all to G␣ q , whereas the RZ and R4 subfamilies tend to interact with both G␣ i/o and G␣ q/11 (10). The exception is RGS2, an R4 subfamily member that is uniquely selective for G␣ q/11 (11). ...

RGS10 Regulates the Expression of Cyclooxygenase-2 and Tumor Necrosis Factor Alpha through a G Protein–Independent Mechanism

“…BV-2 cells were treated with LPS (10 ng/ml) for 3 hours, and then cell lysates were immunoprecipitated with Ga i , Ga q , or control antibodies and the associated proteins were immunoblotted for the G proteins and RGS10. Multiple studies have described strong Ga i selectivity of the RGS10 RGS domain (Ajit and Young, 2005;Taylor et al, 2016); however, it has also been reported that RGS10 can display weak interactions with Ga q (Soundararajan et al, 2008). Our results show that LPS activation of BV-2 cells enhances RGS10 association with Ga i , but no detectable association with Ga q was observed ( Fig.…”

Tyrosine phosphorylation switching of a G protein

“…In animals, RGS proteins typically bind the switch region on their corresponding Gα proteins (60)(61)(62)(63)(64). The reported GAP resistant phosphorylation sites on Gαz were all at N terminus nearby the switch region (15)(16)(17).…”

Atypical activation of the G protein Gαq by the oncogenic mutation Q209P

“…We reasoned that binding of G␣ q Q209P to RGS proteins might be impaired because they also utilize the SwII as an important contact point. In fact, crystal structures of RGS8 and RGS2 in complex with G␣ q (54,55) have revealed that, despite adopting different poses, both RGS proteins make contact with Gln-209 and several adjacent residues in the SwII (Fig. 5, C and D).…”