The association of phospholipase (PLD)-1 with protein kinase C-related prot
ein kinases, PKN alpha and PKN beta, was analyzed. PLD1 interacted with PKN
alpha and PKN beta in COS-7 cells transiently transfected with PLD1 and PK
N alpha or PKN beta expression constructs. The interactions between endogen
ous PLD1 and PKN alpha or PKN beta were confirmed by co-immunoprecipitation
from mammalian cells. In vitro binding studies using the deletion mutants
of PLD1 indicated that PKN alpha directly bound to residues 228-598 of PLD1
and that PKN beta interacted with residues 1-228 and 228-598 of PLD1. PKN
alpha stimulated the activity of PLD1 in the presence of phosphatidylinosit
ol 4,5-bisphosphate in vitro, whereas PKN beta had a modest effect on the s
timulation of PLD1 activity. The stimulation of PLD1 activity by PKN alpha
was slightly enhanced by the addition of arachidonic acid. These results su
ggest that the PKN family functions as a novel intracellular player of PLD1
signaling pathway.