scholarly journals A Rho family GTPase controls actin dynamics and tip growth via two counteracting downstream pathways in pollen tubes

2005 ◽  
Vol 169 (1) ◽  
pp. 127-138 ◽  
Author(s):  
Ying Gu ◽  
Ying Fu ◽  
Peter Dowd ◽  
Shundai Li ◽  
Vanessa Vernoud ◽  
...  
Keyword(s):  
Pollen Tube ◽  
Tip Growth ◽  
Rho Gtpase ◽  
Plant Cells ◽  
Actin Dynamics ◽  
Actin Assembly ◽  
Filamentous Actin ◽  
Growth Defects ◽  
Spatiotemporal Regulation ◽  
Rho Family

Tip growth in neuronal cells, plant cells, and fungal hyphae is known to require tip-localized Rho GTPase, calcium, and filamentous actin (F-actin), but how they interact with each other is unclear. The pollen tube is an exciting model to study spatiotemporal regulation of tip growth and F-actin dynamics. An Arabidopsis thaliana Rho family GTPase, ROP1, controls pollen tube growth by regulating apical F-actin dynamics. This paper shows that ROP1 activates two counteracting pathways involving the direct targets of tip-localized ROP1: RIC3 and RIC4. RIC4 promotes F-actin assembly, whereas RIC3 activates Ca2+ signaling that leads to F-actin disassembly. Overproduction or depletion of either RIC4 or RIC3 causes tip growth defects that are rescued by overproduction or depletion of RIC3 or RIC4, respectively. Thus, ROP1 controls actin dynamics and tip growth through a check and balance between the two pathways. The dual and antagonistic roles of this GTPase may provide a unifying mechanism by which Rho modulates various processes dependent on actin dynamics in eukaryotic cells.

scholarly journals Rho-GTPase–dependent filamentous actin dynamics coordinate vesicle targeting and exocytosis during tip growth

2008 ◽  
Vol 181 (7) ◽  
pp. 1155-1168 ◽  
Author(s):  
Yong Jik Lee ◽  
Amy Szumlanski ◽  
Erik Nielsen ◽  
Zhenbiao Yang
Keyword(s):  
Plasma Membrane ◽  
Tip Growth ◽  
Rho Gtpase ◽  
Actin Dynamics ◽  
Actin Assembly ◽  
Filamentous Actin ◽  
Latrunculin B ◽  
Dynamic Activity ◽  
Guanosine Triphosphatase ◽  
Spatiotemporal Coordination

The dynamic activity of tip-localized filamentous actin (F-actin) in pollen tubes is controlled by counteracting RIC4 and RIC3 pathways downstream of the ROP1 guanosine triphosphatase promoting actin assembly and disassembly, respectively. We show here that ROP1 activation is required for both the polar accumulation and the exocytosis of vesicles at the plasma membrane apex. The apical accumulation of exocytic vesicles oscillated in phase with, but slightly behind, apical actin assembly and was enhanced by overexpression of RIC4. However, RIC4 overexpression inhibited exocytosis, and this inhibition could be suppressed by latrunculin B treatment or RIC3 overexpression. We conclude that RIC4-dependent actin assembly is required for polar vesicle accumulation, whereas RIC3-mediated actin disassembly is required for exocytosis. Thus ROP1-dependent F-actin dynamics control tip growth through spatiotemporal coordination of vesicle targeting and exocytosis.

scholarly journals Grit, a GTPase-Activating Protein for the Rho Family, Regulates Neurite Extension through Association with the TrkA Receptor and N-Shc and CrkL/Crk Adapter Molecules

2002 ◽  
Vol 22 (24) ◽  
pp. 8721-8734 ◽  
Author(s):  
Takeshi Nakamura ◽  
Misako Komiya ◽  
Kiyoaki Sone ◽  
Eiji Hirose ◽  
Noriko Gotoh ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Rho Gtpases ◽  
Rho Gtpase ◽  
Neuronal Cells ◽  
Small Gtpases ◽  
Actin Dynamics ◽  
Gtpase Activating Protein ◽  
Interacting Protein ◽  
High Affinity Receptor ◽  
Rho Family

ABSTRACT Neurotrophins are key regulators of the fate and shape of neuronal cells and act as guidance cues for growth cones by remodeling the actin cytoskeleton. Actin dynamics is controlled by Rho GTPases. We identified a novel Rho GTPase-activating protein (Grit) for Rho/Rac/Cdc42 small GTPases. Grit was abundant in neuronal cells and directly interacted with TrkA, a high-affinity receptor for nerve growth factor (NGF). Another pool of Grit was recruited to the activated receptor tyrosine kinase through its binding to N-Shc and CrkL/Crk, adapter molecules downstream of activated receptor tyrosine kinases. Overexpression of the TrkA-binding region of Grit inhibited NGF-induced neurite elongation. Further, we found some tendency for neurite promotion in full-length Grit-overexpressing PC12 cells upon NGF stimulation. These results suggest that Grit, a novel TrkA-interacting protein, regulates neurite outgrowth by modulating the Rho family of small GTPases.

IgE alone-induced actin assembly modifies calcium signaling and degranulation in RBL-2H3 mast cells

2004 ◽  
Vol 286 (2) ◽  
pp. C256-C263 ◽  
Author(s):  
Tatsuya Oka ◽  
Masatoshi Hori ◽  
Akane Tanaka ◽  
Hiroshi Matsuda ◽  
Hideaki Karaki ◽  
...  
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Cell Activation ◽  
De Novo ◽  
Immunoglobulin E ◽  
Cytochalasin D ◽  
Mast Cell Activation ◽  
Actin Dynamics ◽  
Actin Assembly ◽  
Filamentous Actin

In the mast cell signaling pathways, the binding of immunoglobulin E (IgE) to FcϵRI, its high-affinity receptor, is generally thought to be a passive step. In this study, we examined the effect of IgE alone, that is, without antigen stimulation, on the degranulation in mast cells. Monomeric IgE (500–5,000 ng/ml) alone increased cytosolic Ca2+ level ([Ca2+]i) and induced degranulation in rat basophilic leukemia (RBL)-2H3 mast cells. Monomeric IgE (5,000 ng/ml) alone also increased [Ca2+]i and induced degranulation in bone marrow-derived mast cells. Interestingly, monomeric IgE (5–50 ng/ml) alone, in concentrations too low to induce degranulation, increased filamentous actin content in RBL-2H3 mast cells. We next examined whether actin dynamics affect the IgE alone-induced RBL-2H3 mast cell activation pathways. Cytochalasin D inhibited the ability of IgE alone (50 ng/ml) to induce de novo actin assembly. In cytochalasin D-treated cells, IgE (50 ng/ml) alone increased [Ca2+]i and induced degranulation. We have summarized the current findings into two points. First, IgE alone increases [Ca2+]i and induces degranulation in mast cells. Second, IgE, at concentrations too low to increase either [Ca2+]i or degranulation, significantly induces actin assembly, which serves as a negative feedback control in the mast cell Ca2+ signaling and degranulation.

scholarly journals Rho GTPases and their effector proteins

2000 ◽  
Vol 348 (2) ◽  
pp. 241-255 ◽  
Author(s):  
Anne L. BISHOP ◽  
Alan HALL
Keyword(s):  
Rho Gtpases ◽  
Cell Cycle Progression ◽  
Biological Effects ◽  
Effector Proteins ◽  
Actin Dynamics ◽  
Filamentous Actin ◽  
Cycle Progression ◽  
Cellular Processes ◽  
Specific Effector ◽  
Rho Family

Rho GTPases are molecular switches that regulate many essential cellular processes, including actin dynamics, gene transcription, cell-cycle progression and cell adhesion. About 30 potential effector proteins have been identified that interact with members of the Rho family, but it is still unclear which of these are responsible for the diverse biological effects of Rho GTPases. This review will discuss how Rho GTPases physically interact with, and regulate the activity of, multiple effector proteins and how specific effector proteins contribute to cellular responses. To date most progress has been made in the cytoskeleton field, and several biochemical links have now been established between GTPases and the assembly of filamentous actin. The main focus of this review will be Rho, Rac and Cdc42, the three best characterized mammalian Rho GTPases, though the genetic analysis of Rho GTPases in lower eukaryotes is making increasingly important contributions to this field.

scholarly journals The PAK system links Rho GTPase signaling to thrombin-mediated platelet activation

2013 ◽  
Vol 305 (5) ◽  
pp. C519-C528 ◽  
Author(s):  
Joseph E. Aslan ◽  
Sandra M. Baker ◽  
Cassandra P. Loren ◽  
Kristina M. Haley ◽  
Asako Itakura ◽  
...  
Keyword(s):  
Calcium Signaling ◽  
Platelet Activation ◽  
Rho Gtpase ◽  
Regulatory Protein ◽  
Small Gtpases ◽  
Mass Spectrometry Analysis ◽  
Actin Dynamics ◽  
Nucleotide Exchange ◽  
Spectrometry Analysis ◽  
Rho Family

Regulation of the platelet actin cytoskeleton by the Rho family of small GTPases is essential for the proper maintenance of hemostasis. However, little is known about how intracellular platelet activation from Rho GTPase family members, including Rac, Cdc42, and Rho, translate into changes in platelet actin structures. To better understand how Rho family GTPases coordinate platelet activation, we identified platelet proteins associated with Rac1, a Rho GTPase family member, and actin regulatory protein essential for platelet hemostatic function. Mass spectrometry analysis revealed that upon platelet activation with thrombin, Rac1 associates with a set of effectors of the p21-activated kinases (PAKs), including GIT1, βPIX, and guanine nucleotide exchange factor GEFH1. Platelet activation by thrombin triggered the PAK-dependent phosphorylation of GIT1, GEFH1, and other PAK effectors, including LIMK1 and Merlin. PAK was also required for the thrombin-mediated activation of the MEK/ERK pathway, Akt, calcium signaling, and phosphatidylserine (PS) exposure. Inhibition of PAK signaling prevented thrombin-induced platelet aggregation and blocked platelet focal adhesion and lamellipodia formation in response to thrombin. Together, these results demonstrate that the PAK signaling system is a key orchestrator of platelet actin dynamics, linking Rho GTPase activation downstream of thrombin stimulation to PAK effector function, MAP kinase activation, calcium signaling, and PS exposure in platelets.

scholarly journals Heterodimeric Capping Protein fromArabidopsisIs Regulated by Phosphatidic Acid

2006 ◽  
Vol 17 (4) ◽  
pp. 1946-1958 ◽  
Author(s):  
Shanjin Huang ◽  
Lisa Gao ◽  
Laurent Blanchoin ◽  
Christopher J. Staiger
Keyword(s):  
Pollen Tube ◽  
Phosphatidic Acid ◽  
Actin Polymerization ◽  
Tip Growth ◽  
Pollen Grains ◽  
Actin Binding ◽  
Filamentous Actin ◽  
Suspension Cells ◽  
Capping Protein ◽  
Extracellular Stimuli

The cytoskeleton is a key regulator of morphogenesis, sexual reproduction, and cellular responses to extracellular stimuli. Changes in the cellular architecture are often assumed to require actin-binding proteins as stimulus-response modulators, because many of these proteins are regulated directly by binding to intracellular second messengers or signaling phospholipids. Phosphatidic acid (PA) is gaining widespread acceptance as a major, abundant phospholipid in plants that is required for pollen tube tip growth and mediates responses to osmotic stress, wounding, and phytohormones; however, the number of identified effectors of PA is rather limited. Here we demonstrate that exogenous PA application leads to significant increases in filamentous actin levels in Arabidopsis suspension cells and poppy pollen grains. To investigate further these lipid-induced changes in polymer levels, we analyzed the properties of a key regulator of actin filament polymerization, the heterodimeric capping protein from Arabidopsis thaliana (AtCP). AtCP binds to PA with a Kdvalue of 17 μM and stoichiometry of ∼1:2. It also binds well to PtdIns(4,5)P2, but not to several other phosphoinositide or acidic phospholipids. The interaction with PA inhibited the actin-binding activity of CP. In the presence of PA, CP is unable to block the barbed or rapidly growing and shrinking end of actin filaments. Precapped filament barbed ends can also be uncapped by addition of PA, allowing rapid filament assembly from an actin monomer pool that is buffered with profilin. The findings support a model in which the inhibition of CP activity in cells by elevated PA results in the stimulation of actin polymerization from a large pool of profilin-actin. Such regulation may be important for the response of plant cells to extracellular stimuli as well as for the normal process of pollen tube tip growth.

scholarly journals Rho Family Gtpase Cdc42 Is Essential for the Actin-Based Motility of Shigella in Mammalian Cells

1999 ◽  
Vol 191 (11) ◽  
pp. 1905-1920 ◽  
Author(s):  
Toshihiko Suzuki ◽  
Hitomi Mimuro ◽  
Hiroaki Miki ◽  
Tadaomi Takenawa ◽  
Takuya Sasaki ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Actin Polymerization ◽  
Fluorescent Protein ◽  
Host Cells ◽  
Actin Dynamics ◽  
Cloud Formation ◽  
Actin Assembly ◽  
Egg Extracts ◽  
Rho Family ◽  
Green Fluorescent

Shigella, the causative agent of bacillary dysentery, is capable of directing its movement within host cells by exploiting actin dynamics. The VirG protein expressed at one pole of the bacterium can recruit neural Wiskott-Aldrich syndrome protein (N-WASP), a downstream effector of Cdc42. Here, we show that Cdc42 is required for the actin-based motility of Shigella. Microinjection of a dominant active mutant Cdc42, but not Rac1 or RhoA, into Swiss 3T3 cells accelerated Shigella motility. In add-back experiments in Xenopus egg extracts, addition of a guanine nucleotide dissociation inhibitor for the Rho family, RhoGDI, greatly diminished the bacterial motility or actin assembly, which was restored by adding activated Cdc42. In N-WASP–depleted extracts, the bacterial movement almost arrested was restored by adding exogenous N-WASP but not H208D, an N-WASP mutant defective in binding to Cdc42. In pyrene actin assay, Cdc42 enhanced VirG-stimulating actin polymerization by N-WASP–actin-related protein (Arp)2/3 complex. Actually, Cdc42 stimulated actin cloud formation on the surface of bacteria expressing VirG in a solution containing N-WASP, Arp2/3 complex, and G-actin. Immunohistological study of Shigella-infected cells expressing green fluorescent protein–tagged Cdc42 revealed that Cdc42 accumulated by being colocalized with actin cloud at one pole of intracellular bacterium. Furthermore, overexpression of H208D mutant in cells interfered with the actin assembly of infected Shigella and diminished the intra- and intercellular spreading. These results suggest that Cdc42 activity is involved in initiating actin nucleation mediated by VirG–N-WASP–Arp2/3 complex formed on intracellular Shigella.

scholarly journals Adenovirus E4orf4 Hijacks Rho GTPase-dependent Actin Dynamics to Kill Cells: A Role for Endosome-associated Actin Assembly

2006 ◽  
Vol 17 (7) ◽  
pp. 3329-3344 ◽  
Author(s):  
Amélie Robert ◽  
Nicolas Smadja-Lamère ◽  
Marie-Claude Landry ◽  
Claudia Champagne ◽  
Ryan Petrie ◽  
...  
Keyword(s):  
Rho Gtpases ◽  
Actin Polymerization ◽  
Rho Gtpase ◽  
De Novo ◽  
Strong Support ◽  
Src Family Kinases ◽  
Actin Dynamics ◽  
Death Process ◽  
Actin Assembly ◽  
Orf4 Protein

The adenovirus early region 4 ORF4 protein (E4orf4) triggers a novel death program that bypasses classical apoptotic pathways in human cancer cells. Deregulation of the cell cytoskeleton is a hallmark of E4orf4 killing that relies on Src family kinases and E4orf4 phosphorylation. However, the cytoskeletal targets of E4orf4 and their role in the death process are unknown. Here, we show that E4orf4 translocates to cytoplasmic sites and triggers the assembly of a peculiar juxtanuclear actin–myosin network that drives polarized blebbing and nuclear shrinkage. We found that E4orf4 activates the myosin II motor and triggers de novo actin polymerization in the perinuclear region, promoting endosomes recruitment to the sites of actin assembly. E4orf4-induced actin dynamics requires interaction with Src family kinases and involves a spatial regulation of the Rho GTPases pathways Cdc42/N-Wasp, RhoA/Rho kinase, and Rac1, which make distinct contributions. Remarkably, activation of the Rho GTPases is required for induction of apoptotic-like cell death. Furthermore, inhibition of actin dynamics per se dramatically impairs E4orf4 killing. This work provides strong support for a causal role for endosome-associated actin dynamics in E4orf4 killing and in the regulation of cancer cell fate.

scholarly journals ROP Gtpase–Dependent Dynamics of Tip-Localized F-Actin Controls Tip Growth in Pollen Tubes

2001 ◽  
Vol 152 (5) ◽  
pp. 1019-1032 ◽  
Author(s):  
Ying Fu ◽  
Guang Wu ◽  
Zhenbiao Yang
Keyword(s):  
Tip Growth ◽  
Fluorescent Protein ◽  
Rho Gtpase ◽  
Pollen Tubes ◽  
Actin Binding ◽  
Apical Region ◽  
Polar Growth ◽  
Actin Organization ◽  
Rop Gtpase ◽  
Rho Family

Tip-growing pollen tubes provide a useful model system to study polar growth. Although roles for tip-focused calcium gradient and tip-localized Rho-family GTPase in pollen tube growth is established, the existence and function of tip-localized F-actin have been controversial. Using the green fluorescent protein–tagged actin-binding domain of mouse talin, we found a dynamic form of tip-localized F-actin in tobacco pollen tubes, termed short actin bundles (SABs). The dynamics of SABs during polar growth in pollen tubes is regulated by Rop1At, a Rop GTPase belonging to the Rho family. When overexpressed, Rop1At transformed SAB into a network of fine filaments and induced a transverse actin band behind the tip, leading to depolarized growth. These changes were due to ectopic Rop1At localization to the apical region of the plasma membrane and were suppressed by guanine dissociation inhibitor overexpression, which removed ectopically localized Rop1At. Rop GTPase–activating protein (RopGAP1) overexpression, or Latrunculin B treatments, also recovered normal actin organization and tip growth in Rop1At-overexpressing tubes. Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth. Finally, SAB oscillates and appears at the tip before growth. Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants.

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