Stimulation of actin stress fibre formation mediated by activation of phospholipase D.
BACKGROUND
Agonist-stimulated phospholipase D (PLD) catalyzes the hydrolysis of phosphatidylcholine, generating the putative messenger phosphatidate (PA). Proposed functions for PA, and hence for PLD, include kinase activation, the regulation of small molecular weight GTP-binding proteins, actin polymerization and secretion. It has not been possible to define a physiological function for PLD activation as it is generally stimulated together with other signalling pathways, such as those involving phospholipases A2 and C, phosphatidylinositide (PI) 3-kinase and the p21(ras)/mitogen-activated protein (MAP) kinase cascade.


RESULTS
We report that, in porcine aortic endothelial (PAE) cells, lysophosphatidic acid (LPA) stimulated PLD activity and rapidly generated PA in the absence of other phospholipase, PI 3-kinase or MAP kinase activities. PLD activation was controlled by a tyrosine kinase-regulated pathway. LPA also stimulated actin stress fibre formation, but was inhibited by butan-1-ol; the alcohol also reduced the accumulation of PA. The addition of PA to cells did not stimulate PLD activity, but did cause stress fibre formation in a manner that was insensitive to butan-1-ol. Stimulation of stress fibre formation by LPA and PA was sensitive to genistein, and was inhibited by micro-injection of the Rho-inhibiting C3 exotoxin into PAE cells.


CONCLUSIONS
This study provides the first clear demonstration of a physiological role for PLD activity. In PAE cells, the stimulation of actin stress fibre formation was a consequence of PA generation and, therefore, PLD activation. The results suggest that PA generation is upstream of Rho activation, and imply a role for PLD in the regulation of Rho-mediated pathways.
Cross
M J
MJ
Institute for Cancer Studies, University of Birmingham Medical School, Edgbaston, Birmingham B15 2TH, UK.
Roberts
S
S
Ridley
A J
AJ
Hodgkin
M N
MN
Stewart
A
A
Claesson-Welsh
L
L
Wakelam
M J
MJ
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology; D010739:Phospholipase D / Q000378:metabolism*; D015398:Signal Transduction; D013552:Swine
D000199:Actins; D008246:Lysophospholipids; D010739:Phospholipase D; D019204:GTP-Binding Proteins