Activated RhoA Is a Positive Feedback Regulator of the Lbc Family of Rho Guanine Nucleotide Exchange Factor Proteins

Medina, Frank; Carter, Angela M.; Dada, Olugbenga; Gutowski, Stephen; Hadas, Jana; Chen, James Z.; Sternweis, Paul C.

doi:10.1074/jbc.m113.450056

Cited by 49 publications

(77 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have shown previously that the GEF activity of PDZRhoGEF and its homologs can be facilitated by membrane recruitment to membrane-delimited substrates such as RhoA (27) and that this functional recruitment can mediate hormone signaling in cells (28). In the reconstituted system, GEFs were recruited to the surface of lipid vesicles that contained regulatory partners and substrates tethered to the membrane surface through interaction of polyhistidine tags with Ni-NTA-conjugated lipids.…”

Section: Both Binding Interfaces Contribute To Recruitment Of Pdz-rhomentioning

confidence: 99%

Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ–RhoGEF

Pascoe

Gutowski

Chen

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

PDZ domains are abundant protein interaction modules and typically recognize a short motif at the C terminus of their ligands, with a few residues in the motif endowing the binding specificity. The sequence-based rules, however, cannot fully account for the specificity between the vast number of PDZ domains and ligands in the cell. Plexins are transmembrane receptors that regulate processes such as axon guidance and angiogenesis. Two related guanine nucleotide exchange factors (GEFs), PDZ-RhoGEF and leukemiaassociated RhoGEF (LARG), use their PDZ domains to bind class B plexins and play critical roles in signaling. Here, we present the crystal structure of the full-length cytoplasmic region of PlexinB2 in complex with the PDZ domain of PDZ-RhoGEF. The structure reveals that, in addition to the canonical C-terminal motif/PDZ interaction, the 3D domain of PlexinB2 forms a secondary interface with the PDZ domain. Our biophysical and cell-based assays show that the secondary interface contributes to the specific interaction between plexin and PDZ-RhoGEF and to signaling by plexin in the cell. Formation of secondary interfaces may be a general mechanism for increasing affinity and specificity of modular domain-mediated interactions.PDZ | plexin | signaling | protein interaction module | specificity P lexins are cell surface receptors for semaphorins, extracellular cues that control essential processes such as neuronal axon guidance and vasculature development (1). Binding of semaphorin to the extracellular region of plexin induces formation of the active dimer of the cytoplasmic region, which transduces signal to downstream pathways (2-7). The plexin cytoplasmic region contains a juxtamembrane segment (JM-segment), a RhoGTPase binding domain (RBD), and a GTPase activating protein (GAP) domain (8-10). The GAP domain, activated by the dimerization, transduces signal through converting its substrate GTPase Rap from the GTP-bound active to the GDP-bound inactive state (2, 3). The RBD regulates plexin activity in response to binding of Rho family GTPases, such as Rac1 (reviewed in ref. 11).In addition to the common signaling pathways through the domains shared by all plexins, class B plexins (B1, B2, and B3) mediate a pathway through their unique C terminus. The conserved "VTDL" motif at the C terminus of these plexins binds to the N-terminal PDZ (PSD-95/Discs-large/ZO-1) domains of two related guanine nucleotide exchange factors (GEFs), PDZ-RhoGEF, and leukemia-associated RhoGEF (LARG) (12)(13)(14)(15)(16)(17). This interaction recruits PDZ-RhoGEF and LARG to the plasma membrane, where they promote the exchange of GDP for GTP on RhoA. GTP-bound RhoA binds its downstream effectors and contributes to plexin signaling (13)(14)(15)18). A recent study has shown that deletion of the C terminus of PlexinB2 causes defects in the development of the liver vasculature in mice, highlighting the critical role of the PDZ-RhoGEF/LARG-RhoA pathway in plexin function in vivo (19).More than 250 PDZ domains exist in the human proteome, con...

show abstract

Section: Both Binding Interfaces Contribute To Recruitment Of Pdz-rhomentioning

confidence: 99%

Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ–RhoGEF

Pascoe

Gutowski

Chen

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

show abstract

“…The PH domains of other RhoGEFs, from the LBC family, directly bind to RhoAGTP, which may increase activity by targeting the GEF to a membrane containing RhoAGDP, thereby acting in a positive feedback mechanism, which can be considered allosteric (binding to the PH domain affects activity of the associated catalytic domain). 97 Binding of both phosphoinositide and Arf6GTP has been reported to recruit Grp1 and ARNO to the plasma membrane, 6 where it may encounter the substrates Arf1GDP and Arf6GDP, which could be considered another allosteric behavior as defined as binding of ligand to one site, the PH domain, affecting activity of a second site, the Sec7 domain. The PH domain of FAPP1 functions similarly to specifically target FAPP1 to the Golgi apparatus by binding a lipid (PI4P) and protein (Arf1GTP) simultaneously.…”

Section: -86mentioning

confidence: 99%

Allosteric properties of PH domains in Arf regulatory proteins

et al. 2016

View full text Add to dashboard Cite

Pleckstrin Homology (PH) domains bind phospholipids and proteins. They are critical regulatory elements of a number enzymes including guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) for Ras-superfamily guanine nucleotide binding proteins such as ADP-ribosylation factors (Arfs). Recent studies have indicated that many PH domains may bind more than one ligand cooperatively. Here we discuss the molecular basis of PH domain-dependent allosteric behavior of 2 ADPribosylation factor exchange factors, Grp1 and Brag2, cooperative binding of ligands to the PH domains of Grp1 and the Arf GTPase-activating protein, ASAP1, and the consequences for activity of the associated catalytic domains.

show abstract

“…Such mechanism where the membrane acts directly as an allosteric modulator over protein function was documented for RhoGEFs (48,49) whose catalytic activity is enhanced by tethering to phospholipid vesicles and for phospholipase C-␤ (PLC-␤) whose ability to hydrolyze phosphatidylinositol 4,5-bisphosphate requires electrostatic displacement of the inhibitory "cap" by charged lipid heads of the membrane (50). Similarly, the results of our study show that the primary role in modulating the activity of RGS/G␤5 complexes belongs to the plasma membrane.…”

Section: Discussionmentioning

confidence: 99%

Association with the Plasma Membrane Is Sufficient for Potentiating Catalytic Activity of Regulators of G Protein Signaling (RGS) Proteins of the R7 Subfamily

Muntean

Martemyanov

2016

Journal of Biological Chemistry

View full text Add to dashboard Cite

Regulators of G protein Signaling (RGS) promote deactivation of heterotrimeric G proteins thus controlling the magnitude and kinetics of responses mediated by G protein-coupled receptors (GPCR).In the nervous system, RGS7 and RGS9 -2 play essential role in vision, reward processing, and movement control. Both RGS7 and RGS9 -2 belong to the R7 subfamily of RGS proteins that form macromolecular complexes with R7-binding protein (R7BP). R7BP targets RGS proteins to the plasma membrane and augments their GTPase-accelerating protein (GAP) activity, ultimately accelerating deactivation of G protein signaling. However, it remains unclear if R7BP serves exclusively as a membrane anchoring subunit or further modulates RGS proteins to increase their GAP activity. To directly answer this question, we utilized a rapidly reversible chemically induced protein dimerization system that enabled us to control RGS localization independent from R7BP in living cells. To monitor kinetics of G␣ deactivation, we coupled this strategy with measuring changes in the GAP activity by bioluminescence resonance energy transfer-based assay in a cellular system containing -opioid receptor. This approach was used to correlate changes in RGS localization and activity in the presence or absence of R7BP. Strikingly, we observed that RGS activity is augmented by membrane recruitment, in an orientation independent manner with no additional contributions provided by R7BP. These findings argue that the association of R7 RGS proteins with the membrane environment provides a major direct contribution to modulation of their GAP activity.In the nervous system, signaling through transmembrane G protein-coupled receptors (GPCRs) 2 plays a crucial role in a number of fundamental processes including differentiation, neurotransmission, and synaptic plasticity (1). Upon binding to an extracellular ligand, GPCRs undergo a conformational change that leads to activation of intracellular heterotrimeric G␣␤␥ proteins. This process involves dissociation of G proteins into G␣GTP and G␤␥ subunits freeing them for the interaction with downstream effectors (2). The magnitude of the response depends on the amount of time G proteins spend in their activated state. Therefore, timely deactivation of G proteins is crucial for determining the overall extent of signaling.Deactivation of G proteins is controlled by Regulators of G protein signaling (RGS) proteins which act as GTPase-activating proteins (GAPs) accelerating the rate of GTP hydrolysis on the G␣ subunit thereby promoting return of G protein to its inactive G␣␤␥ heterotrimeric state. It is currently well accepted that the RGS-catalyzed G protein deactivation is essential for determining timing, extent and sensitivity of G protein signaling in a number of GPCR pathways (3, 4). Among more than 30 known RGS proteins in mammalian genomes, critical roles for the signaling in the nervous system has been attributed to the R7 family that includes RGS6, RGS7, RGS9, and RGS11. Two members of the R7 family in particular, RGS7 an...

show abstract

Activated RhoA Is a Positive Feedback Regulator of the Lbc Family of Rho Guanine Nucleotide Exchange Factor Proteins

Cited by 49 publications

References 20 publications

Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ–RhoGEF

Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ–RhoGEF

Allosteric properties of PH domains in Arf regulatory proteins

Association with the Plasma Membrane Is Sufficient for Potentiating Catalytic Activity of Regulators of G Protein Signaling (RGS) Proteins of the R7 Subfamily

Contact Info

Product

Resources

About