scholarly journals β 1 Pix, a RAC/CDC42 Guanine Nucleotide Exchange Factor (GEF), Mediates Cross-Talk Between Wnt/β-Catenin and G-Protein Coupled Receptor (GPCR) Signaling in Colon Cancer

2011 ◽  
Vol 140 (5) ◽  
pp. S-629
Author(s):  
Ahmed Chahdi ◽  
Jean-Pierre Raufman
Keyword(s):  
Colon Cancer ◽  
G Protein ◽  
Cross Talk ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Gpcr Signaling ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
G Protein Coupled

scholarly journals The guanine nucleotide exchange factor Ric-8A induces domain separation and Ras domain plasticity in Gαi1

2015 ◽  
Vol 112 (5) ◽  
pp. 1404-1409 ◽  
Author(s):  
Ned Van Eps ◽  
Celestine J. Thomas ◽  
Wayne L. Hubbell ◽  
Stephen R. Sprang
Keyword(s):  
G Protein ◽  
Bound States ◽  
Guanine Nucleotide Exchange Factor ◽  
Nucleotide Binding ◽  
Conformational Ensemble ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Heterotrimeric G proteins are activated by exchange of GDP for GTP at the G protein alpha subunit (Gα), most notably by G protein-coupled transmembrane receptors. Ric-8A is a soluble cytoplasmic protein essential for embryonic development that acts as both a guanine nucleotide exchange factor (GEF) and a chaperone for Gα subunits of the i, q, and 12/13 classes. Previous studies demonstrated that Ric-8A stabilizes a dynamically disordered state of nucleotide-free Gα as the catalytic intermediate for nucleotide exchange, but no information was obtained on the structures involved or the magnitude of the structural fluctuations. In the present study, site-directed spin labeling (SDSL) together with double electron-electron resonance (DEER) spectroscopy is used to provide global distance constraints that identify discrete members of a conformational ensemble in the Gαi1:Ric-8A complex and the magnitude of structural differences between them. In the complex, the helical and Ras-like nucleotide-binding domains of Gαi1 pivot apart to occupy multiple resolved states with displacements as large as 25 Å. The domain displacement appears to be distinct from that observed in Gαs upon binding of Gs to the β2 adrenergic receptor. Moreover, the Ras-like domain exhibits structural plasticity within and around the nucleotide-binding cavity, and the switch I and switch II regions, which are known to adopt different conformations in the GDP- and GTP-bound states of Gα, undergo structural rearrangements. Collectively, the data show that Ric-8A induces a conformationally heterogeneous state of Gαi and provide insight into the mechanism of action of a nonreceptor Gα GEF.

scholarly journals Ric-8B Is a GTP-dependent G Protein αs Guanine Nucleotide Exchange Factor

2011 ◽  
Vol 286 (22) ◽  
pp. 19932-19942 ◽  
Author(s):  
PuiYee Chan ◽  
Meital Gabay ◽  
Forrest A. Wright ◽  
Gregory G. Tall
Keyword(s):  
G Protein ◽  
Asymmetric Cell Division ◽  
Odorant Receptor ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Gtpγs Binding

ric-8 (resistance to inhibitors of cholinesterase 8) genes have positive roles in variegated G protein signaling pathways, including Gαq and Gαs regulation of neurotransmission, Gαi-dependent mitotic spindle positioning during (asymmetric) cell division, and Gαolf-dependent odorant receptor signaling. Mammalian Ric-8 activities are partitioned between two genes, ric-8A and ric-8B. Ric-8A is a guanine nucleotide exchange factor (GEF) for Gαi/αq/α12/13 subunits. Ric-8B potentiated Gs signaling presumably as a Gαs-class GEF activator, but no demonstration has shown Ric-8B GEF activity. Here, two Ric-8B isoforms were purified and found to be Gα subunit GDP release factor/GEFs. In HeLa cells, full-length Ric-8B (Ric-8BFL) bound endogenously expressed Gαs and lesser amounts of Gαq and Gα13. Ric-8BFL stimulated guanosine 5′-3-O-(thio)triphosphate (GTPγS) binding to these subunits and Gαolf, whereas the Ric-8BΔ9 isoform stimulated Gαs short GTPγS binding only. Michaelis-Menten experiments showed that Ric-8BFL elevated the Vmax of Gαs steady state GTP hydrolysis and the apparent Km values of GTP binding to Gαs from ∼385 nm to an estimated value of ∼42 μm. Directionality of the Ric-8BFL-catalyzed Gαs exchange reaction was GTP-dependent. At sub-Km GTP, Ric-BFL was inhibitory to exchange despite being a rapid GDP release accelerator. Ric-8BFL binds nucleotide-free Gαs tightly, and near-Km GTP levels were required to dissociate the Ric-8B·Gα nucleotide-free intermediate to release free Ric-8B and Gα-GTP. Ric-8BFL-catalyzed nucleotide exchange probably proceeds in the forward direction to produce Gα-GTP in cells.

scholarly journals Crystal and cryo-EM structures of the cytosolic G protein alpha chaperone and guanine nucleotide exchange factor Ric-8A bound to Gαi1

2020 ◽  
Author(s):  
Levi J. McClelland ◽  
Kaiming Zhang ◽  
Tung-Chung Mou ◽  
Jake Johnston ◽  
Cindee Yates-Hansen ◽  
...  
Keyword(s):  
G Protein ◽  
Domain Architecture ◽  
Guanine Nucleotide Exchange Factor ◽  
Chaperone Activity ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Ric-8A is a cytosolic Guanine Nucleotide exchange Factor (GEF) that activates heterotrimeric G protein alpha subunits (Gα)1. Ric-8A is essential to life in multicellular eukaryotes by virtue of its chaperone activity that is required for Gα biogenesis and membrane localization2, 3. Ric-8A adopts an armadillo (ARM)/HEAT repeat domain architecture and is structurally unrelated to G Protein-Coupled Receptors (GPCR)4. Both GEF and chaperone activities are stimulated by Casein Kinase II phosphorylation5. The mechanisms by which Ric-8A catalyzes GDP release and GTP binding to Gα, or exerts chaperone activity are unknown. Here, we report the structure of the nanobody-stabilized complex of nucleotide-free Gαi1 (isoform 1 of Gα family i) and phosphorylated Ric-8A at near atomic resolution by cryo-electron microscopy and X-ray crystallography. We find that Ric-8A envelops the GTPase domain of Gα, disrupting all three switch regions that convey Gα nucleotide-binding and signaling activity, and displaces the C-terminal helix and helical domain of Gα. These cooperative interactions dismantle the GDP binding site and promote GDP release, while protecting structural elements of Gα that are dynamic in the nucleotide-free state. The structures also show how in vivo phosphorylation stabilizes Gα-binding elements of Ric-8A, thereby enhancing its GEF and chaperone activities.

scholarly journals Wsc1 and Mid2 Are Cell Surface Sensors for Cell Wall Integrity Signaling That Act through Rom2, a Guanine Nucleotide Exchange Factor for Rho1

2001 ◽  
Vol 21 (1) ◽  
pp. 271-280 ◽  
Author(s):  
Bevin Philip ◽  
David E. Levin
Keyword(s):  
Cell Wall ◽  
Protein Kinase ◽  
Cell Surface ◽  
G Protein ◽  
Guanine Nucleotide Exchange Factor ◽  
Cell Wall Integrity ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

ABSTRACT Wsc1 and Mid2 are highly O-glycosylated cell surface proteins that reside in the plasma membrane of Saccharomyces cerevisiae. They have been proposed to function as mechanosensors of cell wall stress induced by wall remodeling during vegetative growth and pheromone-induced morphogenesis. These proteins are required for activation of the cell wall integrity signaling pathway that consists of the small G-protein Rho1, protein kinase C (Pkc1), and a mitogen-activated protein kinase cascade. We show here by two-hybrid experiments that the C-terminal cytoplasmic domains of Wsc1 and Mid2 interact with Rom2, a guanine nucleotide exchange factor (GEF) for Rho1. At least with regard to Wsc1, this interaction is mediated by the Rom2 N-terminal domain. This domain is distinct from the Rho1-interacting domain, suggesting that the GEF can interact simultaneously with a sensor and with Rho1. We also demonstrate that extracts from wsc1 and mid2 mutants are deficient in the ability to catalyze GTP loading of Rho1 in vitro, providing evidence that the function of the sensor-Rom2 interaction is to stimulate nucleotide exchange toward this G-protein. In a related line of investigation, we identified the PMT2 gene in a genetic screen for mutations that confer an additive cell lysis defect with a wsc1 null allele. Pmt2 is a member of a six-protein family in yeast that catalyzes the first step in O mannosylation of target proteins. We demonstrate that Mid2 is not mannosylated in apmt2 mutant and that this modification is important for signaling by Mid2.

Sign in / Sign up

Export Citation Format

Share Document