Regulator of G Protein Signaling 7 (RGS7) Can Exist in a Homo-oligomeric Form That Is Regulated by Gαo and R7-binding Protein

Abstract

RGS (regulator of G protein signaling) proteins of the R7 subfamily (RGS6, -7, -9, and -11) are highly expressed in neurons where they regulate many physiological processes. R7 RGS proteins contain several distinct domains and form obligatory dimers with the atypical G␤ subunit, G␤ 5 . They also interact with other proteins such as R7-binding protein, R9-anchoring protein, and the orphan receptors GPR158 and GPR179. These interactions facilitate plasma membrane targeting and stability of R7 proteins and modulate their activity. Here, we investigated RGS7 complexes using in situ chemical cross-linking. We found that in mouse brain and transfected cells cross-linking causes formation of distinct RGS7 complexes. One of the products had the apparent molecular mass of ϳ150 kDa on SDS-PAGE and did not contain G␤ 5 . Mass spectrometry analysis showed no other proteins to be present within the 150-kDa complex in the amount close to stoichiometric with RGS7. This finding suggested that RGS7 could form a homo-oligomer. Indeed, co-immunoprecipitation of differentially tagged RGS7 constructs, with or without chemical cross-linking, demonstrated RGS7 self-association. RGS7-RGS7 interaction required the DEP domain but not the RGS and DHEX domains or the G␤ 5 subunit. Using transfected cells and knock-out mice, we demonstrated that R7-binding protein had a strong inhibitory effect on homooligomerization of RGS7. In contrast, our data indicated that GPR158 could bind to the RGS7 homo-oligomer without causing its dissociation. Co-expression of constitutively active G␣ o prevented the RGS7-RGS7 interaction. These results reveal the existence of RGS protein homo-oligomers and show regulation of their assembly by R7 RGS-binding partners. G protein-coupled receptors (GPCRs)2 regulate many functions in eukaryotic cells by responding to chemically diverse molecules such as biogenic amines, lipids, and peptides. A large fraction of therapeutic drugs act through GPCRs. Although ligand-bound GPCRs directly interact with several proteins, the canonical signaling mechanism involves heterotrimeric (G␣␤␥) G proteins. The receptor promotes the exchange of GDP for GTP on the G␣ subunit and the subsequent dissociation of G␣-GTP from G␤␥. Both G␣-GTP and G␤␥ modulate the activity of effector enzymes or ion channels, thereby causing alterations in second messenger concentrations, which in turn generate cellular responses (1-3).GPCR signaling can be terminated by several mechanisms, one of which involves hydrolysis of the G␣-bound GTP and reassembly of the inactive G␣␤␥ trimer (1). The intrinsic GTPase activity of the G␣ subunit is very slow, and for most G proteins it is accelerated by regulator of G protein signaling (RGS) proteins. The GTPase activating (GAP) function of RGS proteins is mediated by the characteristic ϳ120-amino acid domain ("RGS box") present in all members of the RGS family (4 -8). The role of RGS proteins in the inactivation of GPCR signaling was demonstrated in numerous studies using mice lacking RGS proteins or harboring RG...

Regulation of neurite morphogenesis by interaction between R7 regulator of G protein signaling complexes and G protein subunit Gα13

“…These results are consistent with an independent study in which complexes containing R7BP were detected in lysates of transfected HEK293T cells and mouse brain lysates crosslinked with paraformaldehyde (30).…”

Expression and distribution of trophoblast glycoprotein in the mouse retina

“…To examine the dendritic distribution of TPBG in the mouse OPL, we compared TPBG-CT or TPBG-NT immunoreactivity with that of GPR179 and CtBP2 (Figure 4). GPR179 is localized exclusively to the dendritic tips of ON-bipolar cells, where it is responsible for anchoring the RGS7-Gb5 complex to the postsynaptic membrane (Orhan et al, 2013;Orlandi, Cao, & Martemyanov, 2013;Tayou et al, 2016) in proximity to mGluR6 (Morgans et al, 2007;Ray et al, 2014). TPBG-CT-labeled puncta do not completely overlap GPR179 puncta at the very tips of the RBC dendrites, but do closely associate with them (3C, arrows).…”

DEPDC1/ EEF1A1 Complex Promotes the Progression of Human Osteosarcoma via Downregulation of FOXO3a

“…Genetic changes in EEF1A1 were detected by The Cancer Genome Atlas (TCGA) to explore its potential impact on selected epigenetic modulators. However, the speci c upstream and downstream regulatory molecules that bind to EEF1A1 were not accurately investigated, nor the clinical correlation between the expression of EEF1A1 and the prognosis of tumor patients [28]. Furthermore, the role of EEF1A1 in the proliferation and metastasis of osteosarcoma has not been studied.…”

“…The common feature of this family is the conserved DNA domain, namely, the Forkhead box. This protein family regulates apoptosis, cell cycle, cell proliferation, DNA damage repair, and cancer development, and inhibits tumor cell proliferation [28]. FOXO3a is among the most widely studied members of the Forkhead family.…”