Rac GTPase interacts specifically with phosphatidylinositol 3-kinase

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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Assessment of the Activation of Rho Family GTP-Binding Proteins in Breast Cancer Cells and Specimens

10.21236/ada396849 ◽

2001 ◽

Author(s):

Yi Zheng

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Binding Proteins ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Rho Family

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Activation of the small GTP-binding proteins rho and rac by growth factor receptors

Journal of Cell Science ◽

10.1242/jcs.108.1.225 ◽

1995 ◽

Vol 108 (1) ◽

pp. 225-233 ◽

Cited By ~ 22

Author(s):

C.D. Nobes ◽

P. Hawkins ◽

L. Stephens ◽

A. Hall

Keyword(s):

Plasma Membrane ◽

Growth Factor ◽

Tyrosine Kinase ◽

Phorbol Ester ◽

Lysophosphatidic Acid ◽

Actin Polymerization ◽

Binding Proteins ◽

Platelet Derived Growth Factor ◽

Gtp Binding ◽

Rho Protein

The small GTP-binding proteins, rho and rac, control signal transduction pathways that link growth factor receptors to the activation of actin polymerization. In Swiss 3T3 cells, rho proteins mediate the lysophosphatidic acid and bombesin-induced formation of focal adhesions and actin stress fibres, whilst rac proteins are required for the platelet-derived growth factor-, insulin-, bombesin- and phorbol ester (phorbol 12-myristate 13-acetate)-stimulated actin polymerization at the plasma membrane that results in membrane ruffling. To investigate the role of p85/p110 phosphatidylinositol 3-kinase in the rho and rac signalling pathways, we have used a potent inhibitor of this activity, wortmannin. Wortmannin has no effect on focal adhesion or actin stress fibre formation induced by lysophosphatidic acid, bombesin or microinjected recombinant rho protein. In contrast, it totally inhibits plasma membrane edge-ruffling induced by platelet-derived growth factor and insulin though not by bombesin, phorbol ester or microinjected recombinant rac protein. We conclude that phosphatidylinositol 3,4,5 trisphosphate mediates activation of rac by the platelet-derived growth factor and insulin receptors. The effects of lysophosphatidic acid on the Swiss 3T3 actin cytoskeleton can be blocked by the tyrosine kinase inhibitor, tyrphostin. Since tyrphostin does not inhibit the effects of microinjected rho protein, we conclude that lysophosphatidic acid activation of rho is mediated by a tyrosine kinase.

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Interaction of G protein Gβγ dimers with small GTP-binding proteins of the Rho family

FEBS Letters ◽

10.1016/s0014-5793(96)01327-0 ◽

1996 ◽

Vol 399 (3) ◽

pp. 211-214 ◽

Cited By ~ 24

Author(s):

Rainer Harhammer ◽

Antje Gohla ◽

Günter Schultz

Keyword(s):

G Protein ◽

Binding Proteins ◽

Gtp Binding Proteins ◽

Gtp Binding ◽

Rho Family

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

The Journal of Cell Biology ◽

10.1083/jcb.126.4.1005 ◽

1994 ◽

Vol 126 (4) ◽

pp. 1005-1015 ◽

Cited By ~ 91

Author(s):

J C Norman ◽

L S Price ◽

A J Ridley ◽

A Hall ◽

A Koffer

Keyword(s):

Mast Cells ◽

Actin Filaments ◽

Actin Polymerization ◽

Binding Proteins ◽

Small Gtpases ◽

Secretory Cells ◽

Cortical Region ◽

Permeabilized Cells ◽

Gtp Binding Proteins ◽

Gtp Binding

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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