A role for Rho in receptor- and G protein-stimulated phospholipase C Reduction in phosphatidylinositol 4,5-bisphosphate by Clostridium difficile toxin B

Schmidt, Martina; Bienek, Christine; Rümenapp, Ulrich; C, Zhang; Lümmen, G.; Kh, Jakobs; Just, Ingo; Aktories, Klaus; Moos, Michael; C, von Eichel-Streiber

doi:10.1007/bf00178707

Cited by 44 publications

(45 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…A physical association between RhoA and PIP5K has been reported (19), and the addition of recombinant RhoA stimulates PIP5K activity in lysates of mouse fibroblasts (20,21). Consistent with the role of Rho in PIP5K activation, inactivation of Rho GTPases by Clostridium difficile toxin B reduces cellular PIP 2 levels by 50 -90% in several tissue culture cell lines (22,23). This results in an inhibition of receptormediated inositol phosphate formation by phospholipase C and PIP 2 -sensitive phospholipase D and also induces profound changes in cell morphology.…”

mentioning

confidence: 55%

Rho and Rho-kinase Mediate Thrombin-induced Phosphatidylinositol 4-Phosphate 5-Kinase Trafficking in Platelets

Yang

Carpenter

Abrams

2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) catalyzes the rate-limiting step in the production of phosphatidylinositol 4,5-bisphosphate (PIP 2 ), a signaling phospholipid that contributes to actin dynamics. We have shown in transfected tissue culture cells that PIP5K translocates from the cytosol to the plasma membrane following agonist-induced stimulation of Rho family GTPases. Nonetheless, it is unclear whether Rho GTPases induce PIP5K relocalization in platelets. We used PIP5K isoform-specific immunoblotting and lipid kinase assays to examine the intracellular localization of PIP5K in resting and activated platelets. Using differential centrifugation to separate the membrane skeleton, actin filaments and associated proteins, and cytoplasmic fractions, we found that PIP5K isoforms were translocated from cytosol to actin-rich fractions following stimulation of the thrombin receptor. PIP5K translocation was detectable within 30 s of stimulation and was complete by 2-5 min. This agonist-induced relocalization and activation of PIP5K was inhibited by 8-(4-parachlorophenylthio)-cAMP, a cAMP analogue that inhibits Rho and Rac. In contrast, 8-(4-parachlorophenylthio)-cGMP, a cGMP analogue that inhibits Rac but not Rho, did not affect PIP5K translocation and activation. This suggests that Rho GTPase may be an essential regulator of PIP5K in platelets. Consistent with this hypothesis, we found that C3 exotoxin (a Rho-specific inhibitor) and HA1077 (an inhibitor of the Rho effector, Rho-kinase) also eliminated PIP5K activation and trafficking into the membrane cytoskeleton. Thus, these data indicate that Rho GTPase and its effector Rho-kinase have an intimate relationship with the trafficking and activation of platelet PIP5K. Moreover, these data suggest that relocalization of platelet PIP5K following agonist stimulation may play an important role in regulating the assembly of the platelet cytoskeleton.

show abstract

mentioning

confidence: 55%

Rho and Rho-kinase Mediate Thrombin-induced Phosphatidylinositol 4-Phosphate 5-Kinase Trafficking in Platelets

Yang

Carpenter

Abrams

2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…HEK-293 cell phosphoinositide levels were determined as described previously [25,26]. Briefly, phosphoinositides were labelled by incubating nearly confluent monolayers of cells for 20-24 h with [$H]inositol (0.5-1 µCi\ml) in inositol-free growth medium.…”

Section: Phosphoinositide Analysismentioning

confidence: 99%

“…Since PtdIns(4,5)P # synthesis in HEK-293 cells is apparently controlled by both tyrosine phosphorylation and Rho proteins [25,26], the relationship between these two regulatory mechanisms was examined. Therefore we studied the effects of Rho protein inactivation by C. difficile toxin B on both the genisteininduced fall and the pervanadate-induced rise in phosphoinositide levels.…”

Section: Role Of Rho Proteins In Tyrosine-phosphorylation-dependent Pmentioning

confidence: 99%

“…Studies on the mechanisms of PLD inhibition by Clostridium difficile toxin B and C. botulinum C3 exoenzyme, which inactivate Rho family GTPases [24], indicated that these toxins decrease the cellular level of PtdIns(4,5)P # [25,26]. As receptor stimulation of PLD activity in HEK-293 cells additionally involves a tyrosine-kinase-dependent mechanism [27], as reported similarly for other cell types [28,29], we have examined in this study whether the cellular level of PtdIns(4,5)P # is under the control of tyrosine phosphorylation.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Tyrosine-phosphorylation-dependent and Rho-protein-mediated control of cellular phosphatidylinositol 4,5-bisphosphate levels

Rümenapp

Schmidt

Olesch

et al. 1998

Biochemical Journal

Self Cite

View full text Add to dashboard Cite

The polyphosphoinositide PtdIns(4,5)P # , best known as a substrate for phospholipase C isozymes, has recently been recognized to be involved in a variety of other cellular processes. The aim of this study was to examine whether the cellular levels of this versatile phospholipid are controlled by tyrosine phosphorylation. The studies were performed in human embryonic kidney (HEK)-293 cells stably expressing the M $ muscarinic acetylcholine receptor. Inhibition of tyrosine phosphatases by pervanadate induced an up-to-approx.-2.5-fold increase in the total cellular level of PtdIns(4,5)P # , which was both time-and concentration-dependent. In contrast, the tyrosine kinase inhibitors, genistein and tyrphostin 23, caused a rapid and specific fall in the cellular PtdIns(4,5)P # level and prevented the stimulatory effect of pervanadate on PtdIns(4,5)P # formation. Inactivation of Rho proteins by Clostridium difficile toxin B caused a similar fall in

show abstract

“…Thus, inactivation of Rho GTPases by Clostridium difficile toxin B reduced cellular PIP 2 levels, resulting in inhibition of receptor-mediated inositol phosphate formation by phospholipase C and PIP 2 -sensitive phospholipase D (PLD) (31,32). RhoA (33) and Rac1 (34) have been shown to stimulate PIP 2 synthesis, and PIP5K isoforms appear to function downstream of RhoA and Rac1 in actin organization (15,35).…”

mentioning

confidence: 99%

Activation of Type I Phosphatidylinositol 4-Phosphate 5-Kinase Isoforms by the Rho GTPases, RhoA, Rac1, and Cdc42

Weernink

Meletiadis

Hommeltenberg

et al. 2004

Journal of Biological Chemistry

Self Cite

158

139

View full text Add to dashboard Cite

Type I phosphatidylinositol 4-phosphate 5-kinase (PIP5K) catalyzes the formation of the phospholipid, phosphatidylinositol 4,5-bisphosphate (PIP 2 ), which is implicated in many cellular processes. The Rho GTPases, RhoA and Rac1, have been shown previously to activate PIP5K and to bind PIP5K. Three type I PIP5K isoforms (I␣, I␤, and I␥) have been identified; however, it is unclear whether these isoforms are differentially or even sequentially regulated by Rho GTPases. Here we show that RhoA and Rac1, as well as Cdc42, but not the Ras-like GTPases, RalA and Rap1A, markedly stimulate PIP 2 synthesis by all three PIP5K isoforms expressed in human embryonic kidney 293 cells, both in vitro and in vivo. RhoA-stimulated PIP 2 synthesis by the PIP5K isoforms was mediated by the RhoA effector, Rho-kinase. Stimulation of PIP5K isoforms by Rac1 and Cdc42 was apparently independent of and additive with RhoA-and Rho-kinase, as shown by studies with C3 transferase and Rho-kinase mutants. RhoA, and to a lesser extent Rac1, but not Cdc42, interacted in a nucleotide-independent form with all three PIP5K isoforms. Binding of PIP5K isoforms to GTP-bound, but not GDP-bound, RhoA could be displaced by Rho-kinase, suggesting a direct and constitutive PIP5K-Rho GTPase binding, which, however, does not trigger PIP5K activation. In summary, our findings indicate that synthesis of PIP 2 by the three PIP5K isoforms is controlled by RhoA, acting via Rho-kinase, as well as Rac1 and Cdc42, implicating that regulation of PIP 2 synthesis has a central position in signaling by these three Rho GTPases.Synthesis and turnover of the membrane phospholipid, phosphatidylinositol 4,5-bisphosphate (PIP 2 ),

show abstract

A role for Rho in receptor- and G protein-stimulated phospholipase C Reduction in phosphatidylinositol 4,5-bisphosphate by Clostridium difficile toxin B

Cited by 44 publications

References 28 publications

Rho and Rho-kinase Mediate Thrombin-induced Phosphatidylinositol 4-Phosphate 5-Kinase Trafficking in Platelets

Rho and Rho-kinase Mediate Thrombin-induced Phosphatidylinositol 4-Phosphate 5-Kinase Trafficking in Platelets

Tyrosine-phosphorylation-dependent and Rho-protein-mediated control of cellular phosphatidylinositol 4,5-bisphosphate levels

Activation of Type I Phosphatidylinositol 4-Phosphate 5-Kinase Isoforms by the Rho GTPases, RhoA, Rac1, and Cdc42

Contact Info

Product

Resources

About