Dual requirement for rho and protein kinase C in direct activation of phospholipase D1 through G protein-coupled receptor signaling.
G protein-coupled and tyrosine kinase receptor activation of phospholipase D1 (PLD1) play key roles in agonist-stimulated cellular responses such as regulated exocytosis, actin stress fiber formation, and alterations in cell morphology and motility. Protein Kinase C, ADP-ribosylation factor (ARF), and Rho family members activate PLD1 in vitro; however, the actions of the stimulators on PLD1 in vivo have been proposed to take place through indirect pathways. We have used the yeast split-hybrid system to generate PLD1 alleles that fail to bind to or to be activated by RhoA but that retain wild-type responses to ARF and PKC. These alleles then were employed in combination with alleles unresponsive to PKC or to both stimulators to examine the activation of PLD1 by G protein-coupled receptors. Our results demonstrate that direct stimulation of PLD1 in vivo by RhoA (and by PKC) is critical for significant PLD1 activation but that PLD1 subcellular localization and regulated phosphorylation occur independently of these stimulatory pathways.
Du
G
G
Department of Pharmacology, University Medical Center at Stony Brook, Stony Brook, New York 11794-5140, USA.
Altshuller
Y M
YM
Kim
Y
Y
Han
J M
JM
Ryu
S H
SH
Morris
A J
AJ
Frohman
M A
MA
D020823:ADP-Ribosylation Factor 1 / Q000502:physiology; D000483:Alleles; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D007425:Intracellular Membranes / Q000201:enzymology; D009154:Mutation; D010739:Phospholipase D / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000378:metabolism; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D020742:rhoA GTP-Binding Protein / Q000502:physiology*
D011956:Receptors, Cell Surface; D011493:Protein Kinase C; D010739:Phospholipase D; C106801:phospholipase D1; D020962:Heterotrimeric GTP-Binding Proteins; D020823:ADP-Ribosylation Factor 1; D020742:rhoA GTP-Binding Protein