scholarly journals Characterization of Galpha13-dependent plasma membrane recruitment of p115RhoGEF

2003 ◽  
Vol 371 (3) ◽  
pp. 709-720 ◽  
Author(s):  
Raja BHATTACHARYYA ◽  
Philip B. WEDEGAERTNER
Keyword(s):  
Plasma Membrane ◽  
G Protein ◽  
Small Gtpase ◽  
Pleckstrin Homology Domain ◽  
Nucleotide Exchange ◽  
Pleckstrin Homology ◽  
Α Subunit ◽  
Membrane Recruitment ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

The Ras homology (Rho) guanine nucleotide exchange factor (GEF), p115RhoGEF, provides a direct link between the G-protein α subunit, α13, and the small GTPase Rho. In the present study, we demonstrate that activated mutants of α13 or α12, but not αq, promote the redistribution of p115RhoGEF from the cytoplasm to the plasma membrane (PM). We also show that the PM translocation of p115RhoGEF is promoted by stimulation of thromboxane A2 receptors. Furthermore, we define domains of p115RhoGEF required for its regulated PM recruitment. The RhoGEF RGS (regulators of G-protein signalling) domain of p115RhoGEF is required for PM recruitment, but it is not sufficient for strong α13-promoted PM recruitment, even though it strongly interacts with activated α13. We also identify the pleckstrin homology domain as essential for α13-mediated PM recruitment. An amino acid substitution of lysine to proline at position 677 in the pleckstrin homology domain of p115RhoGEF inhibits Rho-mediated gene transcription, but this mutation does not affect α13-mediated PM translocation of p115RhoGEF. The results suggest a mechanism whereby multiple signals contribute to regulated PM localization of p115RhoGEF.

scholarly journals Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1

2004 ◽  
Vol 382 (3) ◽  
pp. 857-865 ◽  
Author(s):  
Ian N. FLEMING ◽  
Ian H. BATTY ◽  
Alan R. PRESCOTT ◽  
Alex GRAY ◽  
Gursant S. KULAR ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Inositol Phosphates ◽  
Pleckstrin Homology Domain ◽  
Membrane Association ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Pleckstrin Homology ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Binding of the Rac1-specific guanine-nucleotide-exchange factor, Tiam1, to the plasma membrane requires the N-terminal pleckstrin homology domain. In the present study, we show that membrane-association is mediated by binding of PtdIns(4,5)P2 to the pleckstrin homology domain. Moreover, in 1321N1 astrocytoma cells, translocation of Tiam1 to the cytosol, following receptor-mediated stimulation of PtdIns(4,5)P2 breakdown, correlates with decreased Rac1-GTP levels, indicating that membrane-association is required for GDP/GTP exchange on Rac1. In addition, we show that platelet-derived growth factor activates Rac1 in vivo by increasing PtdIns(3,4,5)P3 concentrations, rather than the closely related lipid, PtdIns(3,4)P2. Finally, the data demonstrate that PtdIns(4,5)P2 and PtdIns(3,4,5)P3 bind to the same pleckstrin homology domain in Tiam1 and that soluble inositol phosphates appear to compete with lipids for this binding. Together, these novel observations provide strong evidence that distinct phosphoinositides regulate different functions of this enzyme, indicating that local concentrations of signalling lipids and the levels of cytosolic inositol phosphates will play crucial roles in determining its activity in vivo.

scholarly journals RalGEF2, a Pleckstrin Homology Domain Containing Guanine Nucleotide Exchange Factor for Ral

2000 ◽  
Vol 275 (38) ◽  
pp. 29761-29766 ◽  
Author(s):  
Kim M. T. de Bruyn ◽  
Johan de Rooij ◽  
Rob M. F. Wolthuis ◽  
Holger Rehmann ◽  
Joep Wesenbeek ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Pleckstrin Homology Domain ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Pleckstrin Homology ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

scholarly journals Activation of Cdc42 GTPase upon CRY2-Induced Cortical Recruitment Is Antagonized by GAPs in Fission Yeast

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 2089 ◽  
Author(s):  
Iker Lamas ◽  
Nathalie Weber ◽  
Sophie G. Martin
Keyword(s):  
Fission Yeast ◽  
Positive Feedback ◽  
Antagonistic Activity ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Cell Architecture

The small GTPase Cdc42 is critical for cell polarization in eukaryotic cells. In rod-shaped fission yeast Schizosaccharomyces pombe cells, active GTP-bound Cdc42 promotes polarized growth at cell poles, while inactive Cdc42-GDP localizes ubiquitously also along cell sides. Zones of Cdc42 activity are maintained by positive feedback amplification involving the formation of a complex between Cdc42-GTP, the scaffold Scd2, and the guanine nucleotide exchange factor (GEF) Scd1, which promotes the activation of more Cdc42. Here, we use the CRY2-CIB1 optogenetic system to recruit and cluster a cytosolic Cdc42 variant at the plasma membrane and show that this leads to its moderate activation also on cell sides. Surprisingly, Scd2, which binds Cdc42-GTP, is still recruited to CRY2-Cdc42 clusters at cell sides in individual deletion of the GEFs Scd1 or Gef1. We show that activated Cdc42 clusters at cell sides are able to recruit Scd1, dependent on the scaffold Scd2. However, Cdc42 activity is not amplified by positive feedback and does not lead to morphogenetic changes, due to antagonistic activity of the GTPase activating protein Rga4. Thus, the cell architecture is robust to moderate activation of Cdc42 at cell sides.

Human p63RhoGEF, a novel RhoA-specific guanine nucleotide exchange factor, is localized in cardiac sarcomere

2002 ◽  
Vol 115 (3) ◽  
pp. 629-640 ◽  
Author(s):  
Michel Souchet ◽  
Elodie Portales-Casamar ◽  
David Mazurais ◽  
Susanne Schmidt ◽  
Isabelle Léger ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpases ◽  
Fiber Formation ◽  
Pleckstrin Homology Domain ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Intact Cells ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

The Rho small GTPases are crucial proteins involved in regulation of signal transduction cascades from extracellular stimuli to cell nucleus and cytoskeleton. It has been reported that these GTPases are directly associated with cardiovascular disorders. In this context, we have searched for novel modulators of Rho GTPases, and here we describe p63RhoGEF a new Db1-like guanine nucleotide exchange factor (GEF). P63RhoGEF encodes a 63 kDa protein containing a Db1 homology domain in tandem with a pleckstrin homology domain and is most closely related to the second Rho GEF domain of Trio. Northern blot and in situ analysis have shown that p63RhoGEF is mainly expressed in heart and brain. In vitro guanine nucleotide exchange assays have shown that p63RhoGEF specifically acts on RhoA. Accordingly, p63RhoGEF expression induces RhoA-dependent stress fiber formation in fibroblasts and in H9C2 cardiac myoblasts. Moreover, we show that p63RhoGEF activation of RhoA in intact cells is dependent on the presence of the PH domain. Using a specific anti-p63RhoGEF antibody, we have detected the p63RhoGEF protein by immunocytochemistry in human heart and brain tissue sections. Confocal microscopy shows that p63RhoGEF is located in the sarcomeric I-band mainly constituted of cardiac sarcomeric actin. Together, these results show that p63RhoGEF is a RhoA-specific GEF that may play a key role in actin cytoskeleton reorganization in different tissues, especially in heart cellular morphology.

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