Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 114, Issue 48, Pages E10319-E10328Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1707992114
Keywords
GPCR; GEF; Girdin; DAPLE; integrin
Categories
Funding
- NIH [R01GM112631, R01GM108733, R01GM098367]
- American Cancer Society [RSG-13-362-01-TBE, IRG-72-001-36]
- Karin Grunebaum Cancer Research Foundation
- Hartwell Foundation
Ask authors/readers for more resources
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 G alpha-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 G alpha i that preclude coupling to every known major binding partner [GPCRs, G beta gamma, 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.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available