Actin filament organization in activated mast cells is regulated by heterotrimeric and small GTP-binding proteins.

Rat peritoneal mast cells, both intact and permeabilized, have been used widely as model secretory cells. GTP-binding proteins and calcium play a major role in controlling their secretory response. Here we have examined changes in the organization of actin filaments in intact mast cells after activation by compound 48/80, and in permeabilized cells after direct activation of GTP-binding proteins by GTP-gamma-S. In both cases, a centripetal redistribution of cellular F-actin was observed: the content of F-actin was reduced in the cortical region and increased in the cell interior. The overall F-actin content was increased. Using permeabilized cells, we show that AIF4-, an activator of heterotrimeric G proteins, induces the disassembly of F-actin at the cortex, while the appearance of actin filaments in the interior of the cell is dependent on two small GTPases, rho and rac. Rho was found to be responsible for de novo actin polymerization, presumably from a membrane-bound monomeric pool, while rac was required for an entrapment of the released cortical filaments. Thus, a heterotrimeric G-protein and the small GTPases, rho and rac, participate in affecting the changes in the actin cytoskeleton observed after activation of mast cells.

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The small GTP-binding proteins, Rac and Rho, regulate cytoskeletal organization and exocytosis in mast cells by parallel pathways.

Molecular Biology of the Cell ◽

10.1091/mbc.7.9.1429 ◽

1996 ◽

Vol 7 (9) ◽

pp. 1429-1442 ◽

Cited By ~ 86

Author(s):

J C Norman ◽

L S Price ◽

A J Ridley ◽

A Koffer

Keyword(s):

Mast Cells ◽

Signaling Pathways ◽

Binding Proteins ◽

Small Gtpases ◽

Cortical Actin ◽

Actin Organization ◽

Gtp Binding Proteins ◽

Upstream Regulator ◽

Gtp Binding ◽

Constitutively Active Mutants

In mast cells, activation of GTP-binding proteins induces centripetal reorganization of actin filaments. This effect is due to disassembly, relocalization, and polymerization of F-actin and is dependent on two small GTPases, Rac and Rho. Activities of Rac and Rho are also essential for the secretory function of mast cells. In response to GTP-gamma-S and/or calcium, only a proportion of permeabilized mast cells is capable of secretory response. Here, we have compared actin organization of secreting and nonsecreting cell populations. We show that the cytoskeletal and secretory responses are strongly correlated, indicating a common upstream regulator of the two functions. The secreting cell population preferentially displays both relocalization and polymerization of actin. However, when actin relocalization or polymerization is inhibited by phalloidin or cytochalasin, respectively, secretion is unaffected. Moreover, the ability of the constitutively active mutants of Rac and Rho to enhance secretion is also unaffected in the presence of cytochalasin. Therefore, Rac and Rho control these two functions by divergent, parallel signaling pathways. Cortical actin disassembly occurs in both secreting and nonsecreting populations and does not, by itself, induce exocytosis. A model for the control of exocytosis is proposed that includes at least four GTP-binding proteins and suggests the presence of both shared and divergent signaling pathways from Rac and Rho.

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Control of exocytosis in adrenal chromaffin cells by GTP-binding proteins studied using permeabilized cells and patch-clamp capacitance measurements

Biochemical Society Transactions ◽

10.1042/bst0220468 ◽

1994 ◽

Vol 22 (2) ◽

pp. 468-472 ◽

Cited By ~ 5

Author(s):

Robert D. Burgoyne ◽

Susan E. Handel ◽

Alan Morgan

Keyword(s):

Patch Clamp ◽

Chromaffin Cells ◽

Binding Proteins ◽

Adrenal Chromaffin Cells ◽

Permeabilized Cells ◽

Capacitance Measurements ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Adrenal Chromaffin

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Expression of small GTPases in the roots and nodules of Medicago truncatula cv. Jemalong

Acta Botanica Croatica ◽

10.2478/botcro-2019-0008 ◽

2019 ◽

Vol 78 (1) ◽

pp. 1-8 ◽

Cited By ~ 1

Author(s):

Abdul Razaque Memon ◽

Christiane Katja Schwager ◽

Karsten Niehaus

Keyword(s):

Medicago Truncatula ◽

Binding Proteins ◽

Polymerase Chain Reaction Analysis ◽

Small Gtpases ◽

Infection Process ◽

Nodule Development ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Differential Expression Pattern

Abstract In this study we used Medicago truncatula, to identify and analyze the expression of small GTP-binding proteins (Arf1, Arl1, Sar1, Rabs, Rop/Rac) and their interacting partners in the infection process in the roots and nodules. A real-time polymerase chain reaction analysis was carried out and our results showed that Arf1 (AtArfB1c-like), MtSar1, AtRabA1e-like, AtRabC1-like, MsRab11-like and AtRop7-like genes were highly expressed in the nodules of rhizobium inoculated plants compared to the non-inoculated ones. On the contrary, AtRabA3 like, AtRab5c and MsRac1-like genes were highly expressed in non-infected nitrogen supplied roots of M. truncatula. Other Rab genes (AtRabA4a, AtRabA4c and AtRabG3a-like genes) were nearly equally expressed in both treatments. Interestingly, RbohB (a respiratory burst NADPH oxidase homologue) was more highly expressed in rhizobium infected than in non-infected roots and nodules. Our data show a differential expression pattern of small GTP-binding proteins in roots and nodules of the plants. This study demonstrates an important role of small GTP-binding proteins in symbiosome biogenesis and root nodule development in legumes.

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Farnesylcysteine analogues inhibit store-regulated Ca2+ entry in human platelets: evidence for involvement of small GTP-binding proteins and actin cytoskeleton

Biochemical Journal ◽

10.1042/bj3470183 ◽

2000 ◽

Vol 347 (1) ◽

pp. 183-192 ◽

Cited By ~ 48

Author(s):

Juan A. ROSADO ◽

Stewart O. SAGE

Keyword(s):

Actin Cytoskeleton ◽

Actin Polymerization ◽

Binding Proteins ◽

Selective Inhibition ◽

Human Platelets ◽

Cytochalasin D ◽

Dependent Manner ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Prenylated Proteins

We have investigated the mechanism of Ca2+ entry into fura-2-loaded human platelets by preventing the prenylation of proteins such as small GTP-binding proteins. The farnesylcysteine analogues farnesylthioacetic acid (FTA) and N-acetyl-S-geranylgeranyl-L-cysteine (AGGC), which are inhibitors of the methylation of prenylated and geranylgeranylated proteins respectively, significantly decreased thrombin-evoked increases in intracellular free Ca2+ concentration ([Ca2+]i) in the presence, but not in the absence, of external Ca2+, suggesting a relatively selective inhibition of Ca2+ entry over internal release. Both these compounds and N-acetyl-S-farnesyl-L-cysteine, which had similar effects to those of FTA, also decreased Ca2+ entry evoked by the depletion of intracellular Ca2+ stores with thapsigargin. The inactive control N-acetyl-S-geranyl-L-cysteine was without effect. Patulin, an inhibitor of prenylation that is inert with respect to methyltransferases, also decreased store-regulated Ca2+ entry. Cytochalasin D, an inhibitor of actin polymerization, significantly decreased store-regulated Ca2+ entry in a time-dependent manner. Both cytochalasin D and the farnesylcysteine analogues FTA and AGGC inhibited actin polymerization; however, when evoking the same extent of decrease in actin filament formation, FTA and AGGC showed greater inhibitory effects on Ca2+ entry, indicating a cytoskeleton-independent component in the regulation of Ca2+ entry by small GTP-binding-protein. These findings suggest that prenylated proteins such as small GTP-binding proteins are involved in store-regulated Ca2+ entry through actin cytoskeleton-dependent and cytoskeleton-independent mechanisms in human platelets.

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Cellular responses regulated by rho-related small GTP-binding proteins

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.1993.0067 ◽

1993 ◽

Vol 340 (1293) ◽

pp. 267-271 ◽

Cited By ~ 44

Keyword(s):

Plasma Membrane ◽

Binding Proteins ◽

Focal Adhesions ◽

Receptor Activation ◽

Small Gtpases ◽

Membrane Receptors ◽

Phagocytic Cells ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Plasma Membrane Receptors

Rho-related proteins are members of the ras superfamily of small GTP-binding proteins. Their function in fibroblasts has been analysed using microinjection of living cells. Rho appears to link plasma membrane receptors to the assembly of focal adhesions and actin stress fibres. The closely related protein rac, on the other hand, links receptors to the polymerization of actin at the plasma membrane to form membrane ruffles and pinocytotic vesicles. In phagocytic cells, rac has been shown to be required for activation of a membrane-bound NADPH oxidase in response to receptor activation. These systems provide the basis for a working model for the mechanism of action of the rho family of small GTPases.

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Rac GTPase interacts specifically with phosphatidylinositol 3-kinase

Biochemical Journal ◽

10.1042/bj3150775 ◽

1996 ◽

Vol 315 (3) ◽

pp. 775-779 ◽

Cited By ~ 134

Author(s):

Gary M. BOKOCH ◽

Chris J. VLAHOS ◽

Yan WANG ◽

Ulla G. KNAUS ◽

Alexis E. TRAYNOR-KAPLAN

Keyword(s):

Growth Factor ◽

Actin Polymerization ◽

Binding Proteins ◽

Specific Interaction ◽

Bound State ◽

Rac Gtpase ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Rho Family

The Rac GTP-binding proteins are members of the Rho family and regulate growth factor-stimulated actin assembly in a variety of cells. The formation of phosphorylated inositol lipids has been implicated in control of the processes initiating and regulating such actin polymerization. Associations of Rho family GTP-binding proteins with enzymes involved in lipid metabolism have been described. Here we demonstrate a direct and specific interaction of Rac proteins with phosphatidylinositol (PI) 3-kinase. This interaction is dependent upon Rac being in a GTP-bound state and requires an intact Rac effector domain. In contrast, direct binding of RhoA to PI 3-kinase could not be detected. Rac–GTP also bound to PI 3-kinase in Swiss 3T3 fibroblast and human neutrophil lysates, and increased PI 3-kinase activity became associated with Rac–GTP in platelet-derived growth factor-stimulated cells. Interaction of Rac–GTP with PI 3-kinase in vitro stimulated the activity of the enzyme by 2–9-fold. A specific interaction of active Rac with PI 3-kinase might be important in regulation of the actin cytoskeleton.

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Endomembrane Trafficking in Plants

Electrodialysis ◽

10.5772/intechopen.91642 ◽

2020 ◽

Author(s):

Birsen Cevher-Keskin

Keyword(s):

Coat Protein ◽

Plant Development ◽

Protein Complex ◽

Binding Proteins ◽

Critical Role ◽

Small Gtpases ◽

Eukaryotic Cells ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Coat Protein Complex

The functional organization of eukaryotic cells requires the exchange of proteins, lipids, and polysaccharides between membrane compartments through transport intermediates. Small GTPases largely control membrane traffic, which is essential for the survival of all eukaryotes. Transport from one compartment of this pathway to another is mediated by vesicular carriers, which are formed by the controlled assembly of coat protein complexes (COPs) on donor organelles. The activation of small GTPases is essential for vesicle formation from a donor membrane. In eukaryotic cells, small GTP-binding proteins comprise the largest family of signaling proteins. The ADP-ribosylation factor 1 (ARF1) and secretion-associated RAS superfamily 1 (SAR1) GTP-binding proteins are involved in the formation and budding of vesicles throughout plant endomembrane systems. ARF1 has been shown to play a critical role in coat protein complex I (COPI)-mediated retrograde trafficking in eukaryotic systems, whereas SAR1 GTPases are involved in intracellular coat protein complex II (COPII)-mediated protein trafficking from the endoplasmic reticulum (ER) to the Golgi apparatus. The dysfunction of the endomembrane system can affect signal transduction, plant development, and defense. This chapter offers a summary of membrane trafficking system with an emphasis on the role of GTPases especially ARF1, SAR1, and RAB, their regulatory proteins, and interaction with endomembrane compartments. The vacuolar and endocytic trafficking are presented to enhance our understanding of plant development and immunity in plants.

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