Kinetic Scaffolding Mediated by a Phospholipase C–β and G
            <sub>q</sub>
            Signaling Complex

Waldo, Gary L.; Ricks, Tiffany K.; Hicks, Stephanie; Cheever, Matthew L.; Kawano, Takeharu; Tsuboi, Kazuhito; Wang, Xiaoyue; Montell, Craig; Kozasa, Tohru; Sondek, John; Harden, T. Kendall

doi:10.1126/science.1193438

Cited by 217 publications

(394 citation statements)

References 39 publications

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“…In contrast, the C-terminal domain is not required for activation of PLC-␤ isozymes by either G␤␥ or Rac1 (20,21). The C-terminal domain was previously thought to interact directly with G␣ q in the activation of PLC-␤ isozymes, but our measurements using soluble proteins indicate that PLC-␤3 lacking the C-terminal domain binds active G␣ q with an affinity only 2-fold lower than that of full-length PLC-␤3 (22). These affinity measurements are supported by a recent crystal structure of full-length PLC-␤3 and G␣ q showing minimal interaction between G␣ q and the C-terminal domain (18).…”

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confidence: 50%

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Charpentier

Waldo

Barrett

et al. 2014

Journal of Biological Chemistry

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Background: Phospholipase C-␤ (PLC-␤) isozymes hydrolyze phosphatidylinositol 4,5-bisphosphate to propagate signals for several physiological responses. Results: Membranes are essential for the allosteric release of autoinhibition of PLC-␤ isozymes. Conclusion: Activators of PLC-␤ release autoinhibition by orientating the isozymes at the membrane. Significance: The model described provides a better understanding of PLC-␤ regulation and potential mechanisms to inhibit their activation.

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confidence: 50%

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Charpentier

Waldo

Barrett

et al. 2014

Journal of Biological Chemistry

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“…The crystal structure of Gα q , which shares 90% amino acid identity with Gα 11 ,3, 20 was used to predict the effects of the Val340Met mutation. The mutated Val340 residue is located within the α5 helix of the GTPase domain of Gα 11 (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The crystal structure of Gα q in complex with the GTP analogue guanosine diphosphate (GDP)‐AIF 4 has been determined (Protein Data Bank [PDB] accession number 3OHM),20 and this was used to model Gα 11 , which has 90% identity with Gα q at the amino acid level, using the PyMOL Molecular Graphics System (version 1.2r3pre; Schrodinger, LLC).…”

Section: Methodsmentioning

confidence: 99%

Identification of a G-Protein Subunit-α11 Gain-of-Function Mutation, Val340Met, in a Family With Autosomal Dominant Hypocalcemia Type 2 (ADH2)

Piret

Gorvin

Pagnamenta

et al. 2016

Journal of Bone and Mineral Research

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Autosomal dominant hypocalcemia (ADH) is characterized by hypocalcemia, inappropriately low serum parathyroid hormone concentrations and hypercalciuria. ADH is genetically heterogeneous with ADH type 1 (ADH1), the predominant form, being caused by germline gain‐of‐function mutations of the G‐protein coupled calcium‐sensing receptor (CaSR), and ADH2 caused by germline gain‐of‐function mutations of G‐protein subunit α‐11 (Gα11). To date Gα11 mutations causing ADH2 have been reported in only five probands. We investigated a multigenerational nonconsanguineous family, from Iran, with ADH and keratoconus which are not known to be associated, for causative mutations by whole‐exome sequencing in two individuals with hypoparathyroidism, of whom one also had keratoconus, followed by cosegregation analysis of variants. This identified a novel heterozygous germline Val340Met Gα11 mutation in both individuals, and this was also present in the other two relatives with hypocalcemia that were tested. Three‐dimensional modeling revealed the Val340Met mutation to likely alter the conformation of the C‐terminal α5 helix, which may affect G‐protein coupled receptor binding and G‐protein activation. In vitro functional expression of wild‐type (Val340) and mutant (Met340) Gα11 proteins in HEK293 cells stably expressing the CaSR, demonstrated that the intracellular calcium responses following stimulation with extracellular calcium, of the mutant Met340 Gα11 led to a leftward shift of the concentration‐response curve with a significantly (p < 0.0001) reduced mean half‐maximal concentration (EC50) value of 2.44 mM (95% CI, 2.31 to 2.77 mM) when compared to the wild‐type EC50 of 3.14 mM (95% CI, 3.03 to 3.26 mM), consistent with a gain‐of‐function mutation. A novel His403Gln variant in transforming growth factor, beta‐induced (TGFBI), that may be causing keratoconus was also identified, indicating likely digenic inheritance of keratoconus and ADH2 in this family. In conclusion, our identification of a novel germline gain‐of‐function Gα11 mutation, Val340Met, causing ADH2 demonstrates the importance of the Gα11 C‐terminal region for G‐protein function and CaSR signal transduction. © 2016 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research (ASBMR).

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“…We utilized these data along with the interaction between G␣ q and its effectors PLC-␤ and GRK2 as models in order to identify specific amino acid residues responsible for activating p115RhoGEF. Recent work has identified several residues that are critical for the interaction between G␣ q and these effectors, namely Ala 253 , Thr 257 , and Tyr 261 (20,33). 3 The G␣ q -PLC-␤ interaction was selected as a model because, like RHRhoGEFs, PLC-␤ functions both as a GAP (34) and effector (35) for G␣ q .…”

Section: Resultsmentioning

confidence: 99%

Identification of Critical Residues in Gα13 for Stimulation of p115RhoGEF Activity and the Structure of the Gα13-p115RhoGEF Regulator of G Protein Signaling Homology (RH) Domain Complex

Hajicek

Kukimoto-Niino

Mishima-Tsumagari

et al. 2011

Journal of Biological Chemistry

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RH-RhoGEFs are a family of guanine nucleotide exchange factors that contain a regulator of G protein signaling homology (RH) domain. The heterotrimeric G protein G␣ 13 stimulates the guanine nucleotide exchange factor (GEF) activity of RHRhoGEFs, leading to activation of RhoA. The mechanism by which G␣ 13 stimulates the GEF activity of RH-RhoGEFs, such as p115RhoGEF, has not yet been fully elucidated. Here, specific residues in G␣ 13 that mediate activation of p115RhoGEF are identified. Mutation of these residues significantly impairs binding of G␣ 13 to p115RhoGEF as well as stimulation of GEF activity. These data suggest that the exchange activity of p115RhoGEF is stimulated allosterically by G␣ 13 and not through its interaction with a secondary binding site. A crystal structure of G␣ 13 bound to the RH domain of p115RhoGEF is also presented, which differs from a previously crystallized complex with a G␣ 13 -G␣ i1 chimera. Taken together, these data provide new insight into the mechanism by which p115RhoGEF is activated by G␣ 13 .Heterotrimeric guanine nucleotide binding proteins (G proteins), composed of ␣, ␤, and ␥ subunits, act as molecular switches that cycle between an inactive, GDP-bound state and an active, GTP-bound state upon stimulation of G protein-coupled receptors (1). Once activated, the GTP-bound G␣ subunit dissociates from the G␤␥ dimer, both of which regulate the activity of multiple intracellular effectors to elicit cellular responses. The duration of signaling mediated by G␣ is dictated by the lifetime of bound GTP. All G␣ subunits have intrinsic GTPase activity that hydrolyzes GTP to GDP, and this rate of hydrolysis is enhanced by GAPs, 2 such as RGS proteins (2), which bind the switch regions of activated G␣ subunits, stabilize the transition state for GTP hydrolysis, and thus accelerate the hydrolysis of GTP to GDP (3, 4). Once in the GDP-bound state, G␣ subunits reassociate with G␤␥ and are capable of initiating another round of signaling.G␣ 13 is one of two members of the G 12 family of G proteins (5) and has been implicated in regulating multiple cellular processes that depend on activation of the monomeric GTPase RhoA, including gene transcription, embryogenesis, and rearrangement of the actin cytoskeleton (6 -8). A direct link between G␣ 13 and RhoA was established with the discovery of p115RhoGEF, the founding member of a family of RhoA-specific GEFs containing an RH domain in their N termini (RHRhoGEFs) (9). Together with p115RhoGEF, PDZ-RhoGEF and LARG constitute the RH-RhoGEF family in mammals (10, 11). The RH domain of p115RhoGEF has low sequence similarity to canonical RGS proteins but nevertheless functions as a GAP for G␣ 13 (12). Although the structure of the RH domain of p115RhoGEF is similar to that of other RGS proteins (13), additional elements flanking the RGS box are required for GAP activity (14) and stability of the isolated domain (15). In addition to its RH domain, p115RhoGEF also contains the tandem DH/PH domains characteristic of Dbl family RhoGEFs. The nucleoti...

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Kinetic Scaffolding Mediated by a Phospholipase C–β and G _q Signaling Complex

Cited by 217 publications

References 39 publications

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Identification of a G-Protein Subunit-α11 Gain-of-Function Mutation, Val340Met, in a Family With Autosomal Dominant Hypocalcemia Type 2 (ADH2)

Identification of Critical Residues in Gα13 for Stimulation of p115RhoGEF Activity and the Structure of the Gα13-p115RhoGEF Regulator of G Protein Signaling Homology (RH) Domain Complex

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Kinetic Scaffolding Mediated by a Phospholipase C–β and G q Signaling Complex

Cited by 217 publications

References 39 publications

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Membrane-induced Allosteric Control of Phospholipase C-β Isozymes

Identification of a G-Protein Subunit-α11 Gain-of-Function Mutation, Val340Met, in a Family With Autosomal Dominant Hypocalcemia Type 2 (ADH2)

Identification of Critical Residues in Gα13 for Stimulation of p115RhoGEF Activity and the Structure of the Gα13-p115RhoGEF Regulator of G Protein Signaling Homology (RH) Domain Complex

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Kinetic Scaffolding Mediated by a Phospholipase C–β and G _q Signaling Complex