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Proc Natl Acad Sci U S A
2017 Nov 28;11448:E10319-E10328. doi: 10.1073/pnas.1707992114.
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Specific inhibition of GPCR-independent G protein signaling by a rationally engineered protein.
Leyme A
,
Marivin A
,
Maziarz M
,
DiGiacomo V
,
Papakonstantinou MP
,
Patel PP
,
Blanco-Canosa JB
,
Walawalkar IA
,
Rodriguez-Davila G
,
Dominguez I
,
Garcia-Marcos M
.
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Activation of heterotrimeric G proteins by cytoplasmic nonreceptor proteins is an alternative to the classical mechanism via G protein-coupled receptors (GPCRs). A subset of nonreceptor G protein activators is characterized by a conserved sequence named the Gα-binding and activating (GBA) motif, which confers guanine nucleotide exchange factor (GEF) activity in vitro and promotes G protein-dependent signaling in cells. GBA proteins have important roles in physiology and disease but remain greatly understudied. This is due, in part, to the lack of efficient tools that specifically disrupt GBA motif function in the context of the large multifunctional proteins in which they are embedded. This hindrance to the study of alternative mechanisms of G protein activation contrasts with the wealth of convenient chemical and genetic tools to manipulate GPCR-dependent activation. Here, we describe the rational design and implementation of a genetically encoded protein that specifically inhibits GBA motifs: GBA inhibitor (GBAi). GBAi was engineered by introducing modifications in Gαi that preclude coupling to every known major binding partner [GPCRs, Gβγ, effectors, guanine nucleotide dissociation inhibitors (GDIs), GTPase-activating proteins (GAPs), or the chaperone/GEF Ric-8A], while favoring high-affinity binding to all known GBA motifs. We demonstrate that GBAi does not interfere with canonical GPCR-G protein signaling but blocks GBA-dependent signaling in cancer cells. Furthermore, by implementing GBAi in vivo, we show that GBA-dependent signaling modulates phenotypes during Xenopus laevis embryonic development. In summary, GBAi is a selective, efficient, and convenient tool to dissect the biological processes controlled by a GPCR-independent mechanism of G protein activation mediated by cytoplasmic factors.
Aznar,
Daple is a novel non-receptor GEF required for trimeric G protein activation in Wnt signaling.
2015, Pubmed
Aznar,
Daple is a novel non-receptor GEF required for trimeric G protein activation in Wnt signaling.
2015,
Pubmed
Berghuis,
Structure of the GDP-Pi complex of Gly203-->Ala gialpha1: a mimic of the ternary product complex of galpha-catalyzed GTP hydrolysis.
1996,
Pubmed
Cabrita,
A family of E. coli expression vectors for laboratory scale and high throughput soluble protein production.
2006,
Pubmed
Charpentier,
Potent and Selective Peptide-based Inhibition of the G Protein Gαq.
2016,
Pubmed
Coleman,
Evolutionary Conservation of a GPCR-Independent Mechanism of Trimeric G Protein Activation.
2016,
Pubmed
De Vries,
RGS-GAIP, a GTPase-activating protein for Galphai heterotrimeric G proteins, is located on clathrin-coated vesicles.
1998,
Pubmed
Denker,
Mutagenesis of the amino terminus of the alpha subunit of the G protein Go. In vitro characterization of alpha o beta gamma interactions.
1992,
Pubmed
DiGiacomo,
The Gαi-GIV binding interface is a druggable protein-protein interaction.
2017,
Pubmed
Ekici,
Disturbed Wnt Signalling due to a Mutation in CCDC88C Causes an Autosomal Recessive Non-Syndromic Hydrocephalus with Medial Diverticulum.
2010,
Pubmed
Enomoto,
Roles of disrupted-in-schizophrenia 1-interacting protein girdin in postnatal development of the dentate gyrus.
2009,
Pubmed
Fischer,
Promotion of G alpha i3 subunit down-regulation by GIPN, a putative E3 ubiquitin ligase that interacts with RGS-GAIP.
2003,
Pubmed
Flock,
Universal allosteric mechanism for Gα activation by GPCRs.
2015,
Pubmed
Garcia-Marcos,
G Protein binding sites on Calnuc (nucleobindin 1) and NUCB2 (nucleobindin 2) define a new class of G(alpha)i-regulatory motifs.
2011,
Pubmed
Garcia-Marcos,
GIV/Girdin transmits signals from multiple receptors by triggering trimeric G protein activation.
2015,
Pubmed
Garcia-Marcos,
Functional characterization of the guanine nucleotide exchange factor (GEF) motif of GIV protein reveals a threshold effect in signaling.
2012,
Pubmed
Garcia-Marcos,
GIV is a nonreceptor GEF for G alpha i with a unique motif that regulates Akt signaling.
2009,
Pubmed
Garcia-Marcos,
A structural determinant that renders G alpha(i) sensitive to activation by GIV/girdin is required to promote cell migration.
2010,
Pubmed
Ghosh,
Heterotrimeric G proteins as emerging targets for network based therapy in cancer: End of a long futile campaign striking heads of a Hydra.
2015,
Pubmed
Gilman,
G proteins: transducers of receptor-generated signals.
1987,
Pubmed
Graf,
A truncated recombinant alpha subunit of Gi3 with a reduced affinity for beta gamma dimers and altered guanosine 5'-3-O-(thio)triphosphate binding.
1992,
Pubmed
Habas,
Wnt/Frizzled activation of Rho regulates vertebrate gastrulation and requires a novel Formin homology protein Daam1.
2001,
Pubmed
,
Xenbase
Hawes,
Distinct pathways of Gi- and Gq-mediated mitogen-activated protein kinase activation.
1995,
Pubmed
Herrmann,
Signal transfer from GPCRs to G proteins: role of the G alpha N-terminal region in rhodopsin-transducin coupling.
2006,
Pubmed
Hollins,
The c-terminus of GRK3 indicates rapid dissociation of G protein heterotrimers.
2009,
Pubmed
Johnston,
Minimal determinants for binding activated G alpha from the structure of a G alpha(i1)-peptide dimer.
2006,
Pubmed
Journot,
Deletion within the amino-terminal region of Gs alpha impairs its ability to interact with beta gamma subunits and to activate adenylate cyclase.
1991,
Pubmed
Kimple,
RGS12 and RGS14 GoLoco motifs are G alpha(i) interaction sites with guanine nucleotide dissociation inhibitor Activity.
2001,
Pubmed
Kimple,
A high throughput fluorescence polarization assay for inhibitors of the GoLoco motif/G-alpha interaction.
2008,
Pubmed
Kitamura,
Regulation of VEGF-mediated angiogenesis by the Akt/PKB substrate Girdin.
2008,
Pubmed
Kobayashi,
Novel Daple-like protein positively regulates both the Wnt/beta-catenin pathway and the Wnt/JNK pathway in Xenopus.
2005,
Pubmed
,
Xenbase
Koch,
Direct evidence that Gi-coupled receptor stimulation of mitogen-activated protein kinase is mediated by G beta gamma activation of p21ras.
1994,
Pubmed
Koch,
Cellular expression of the carboxyl terminus of a G protein-coupled receptor kinase attenuates G beta gamma-mediated signaling.
1994,
Pubmed
Le-Niculescu,
Identification and characterization of GIV, a novel Galpha i/s-interacting protein found on COPI, endoplasmic reticulum-Golgi transport vesicles.
2005,
Pubmed
Leyme,
GIV/Girdin (Gα-interacting, Vesicle-associated Protein/Girdin) Creates a Positive Feedback Loop That Potentiates Outside-in Integrin Signaling in Cancer Cells.
2016,
Pubmed
Leyme,
Integrins activate trimeric G proteins via the nonreceptor protein GIV/Girdin.
2015,
Pubmed
Lin,
Calnuc plays a role in dynamic distribution of Galphai but not Gbeta subunits and modulates ACTH secretion in AtT-20 neuroendocrine secretory cells.
2009,
Pubmed
Lin,
Overexpression of CALNUC (nucleobindin) increases agonist and thapsigargin releasable Ca2+ storage in the Golgi.
1999,
Pubmed
Lin,
Structural basis for activation of trimeric Gi proteins by multiple growth factor receptors via GIV/Girdin.
2014,
Pubmed
Lopez-Sanchez,
Focal adhesions are foci for tyrosine-based signal transduction via GIV/Girdin and G proteins.
2015,
Pubmed
Lopez-Sanchez,
GIV/Girdin is a central hub for profibrogenic signalling networks during liver fibrosis.
2014,
Pubmed
Ma,
Therapeutic effects of cell-permeant peptides that activate G proteins downstream of growth factors.
2015,
Pubmed
Marivin,
Dominant-negative Gα subunits are a mechanism of dysregulated heterotrimeric G protein signaling in human disease.
2016,
Pubmed
,
Xenbase
Masuho,
Distinct profiles of functional discrimination among G proteins determine the actions of G protein-coupled receptors.
2015,
Pubmed
Maziarz,
Fluorescence polarization assays to measure interactions between Gα subunits of heterotrimeric G proteins and regulatory motifs.
2017,
Pubmed
Maziarz,
Rapid kinetic BRET measurements to monitor G protein activation by GPCR and non-GPCR proteins.
2017,
Pubmed
Midde,
Multimodular biosensors reveal a novel platform for activation of G proteins by growth factor receptors.
2015,
Pubmed
Milligan,
Heterotrimeric G-proteins: a short history.
2006,
Pubmed
Nakai,
Girdin phosphorylation is crucial for synaptic plasticity and memory: a potential role in the interaction of BDNF/TrkB/Akt signaling with NMDA receptor.
2014,
Pubmed
Nanoff,
The carboxyl terminus of the Galpha-subunit is the latch for triggered activation of heterotrimeric G proteins.
2006,
Pubmed
Neer,
The amino terminus of G protein alpha subunits is required for interaction with beta gamma.
1988,
Pubmed
Oldham,
Mechanism of the receptor-catalyzed activation of heterotrimeric G proteins.
2006,
Pubmed
Oshita,
Identification and characterization of a novel Dvl-binding protein that suppresses Wnt signalling pathway.
2003,
Pubmed
,
Xenbase
Parag-Sharma,
Membrane Recruitment of the Non-receptor Protein GIV/Girdin (Gα-interacting, Vesicle-associated Protein/Girdin) Is Sufficient for Activating Heterotrimeric G Protein Signaling.
2016,
Pubmed
Park,
Subcellular localization and signaling properties of dishevelled in developing vertebrate embryos.
2005,
Pubmed
,
Xenbase
Rasmussen,
Crystal structure of the β2 adrenergic receptor-Gs protein complex.
2011,
Pubmed
Raw,
Structural and biochemical characterization of the GTPgammaS-, GDP.Pi-, and GDP-bound forms of a GTPase-deficient Gly42 --> Val mutant of Gialpha1.
1997,
Pubmed
Ross,
GTPase-activating proteins for heterotrimeric G proteins: regulators of G protein signaling (RGS) and RGS-like proteins.
2000,
Pubmed
Sato,
Accessory proteins for G proteins: partners in signaling.
2006,
Pubmed
Schumacher,
A peptide of the RGS domain of GRK2 binds and inhibits Gα(q) to suppress pathological cardiac hypertrophy and dysfunction.
2016,
Pubmed
Sjögren,
Regulators of G protein signaling proteins as targets for drug discovery.
2010,
Pubmed
Smrcka,
Molecular targeting of Gα and Gβγ subunits: a potential approach for cancer therapeutics.
2013,
Pubmed
Sokol,
Analysis of Dishevelled signalling pathways during Xenopus development.
1996,
Pubmed
,
Xenbase
Stols,
A new vector for high-throughput, ligation-independent cloning encoding a tobacco etch virus protease cleavage site.
2002,
Pubmed
Sun,
Probing Gαi1 protein activation at single-amino acid resolution.
2015,
Pubmed
Tada,
Xwnt11 is a target of Xenopus Brachyury: regulation of gastrulation movements via Dishevelled, but not through the canonical Wnt pathway.
2000,
Pubmed
,
Xenbase
Tall,
Ric-8 regulation of heterotrimeric G proteins.
2013,
Pubmed
Tesmer,
Structure of RGS4 bound to AlF4--activated G(i alpha1): stabilization of the transition state for GTP hydrolysis.
1997,
Pubmed
Thomas,
The nucleotide exchange factor Ric-8A is a chaperone for the conformationally dynamic nucleotide-free state of Gαi1.
2011,
Pubmed
Thomas,
Uncoupling conformational change from GTP hydrolysis in a heterotrimeric G protein alpha-subunit.
2004,
Pubmed
Thomas,
Ric-8A catalyzes guanine nucleotide exchange on G alphai1 bound to the GPR/GoLoco exchange inhibitor AGS3.
2008,
Pubmed
Waldo,
Kinetic scaffolding mediated by a phospholipase C-beta and Gq signaling complex.
2010,
Pubmed
Wall,
Structural basis of activity and subunit recognition in G protein heterotrimers.
1998,
Pubmed
Wallingford,
Convergent extension: the molecular control of polarized cell movement during embryonic development.
2002,
Pubmed
,
Xenbase
Weiss,
Galpha i3 binding to calnuc on Golgi membranes in living cells monitored by fluorescence resonance energy transfer of green fluorescent protein fusion proteins.
2001,
Pubmed
Willard,
A point mutation to Galphai selectively blocks GoLoco motif binding: direct evidence for Galpha.GoLoco complexes in mitotic spindle dynamics.
2008,
Pubmed
Willard,
Return of the GDI: the GoLoco motif in cell division.
2004,
Pubmed
de Opakua,
Molecular mechanism of Gαi activation by non-GPCR proteins with a Gα-Binding and Activating motif.
2017,
Pubmed
