Submitted February 27, 2006
Accepted June 24, 2006
Phagocyte cell migration is mediated by phospholipases PLD1 and PLD2
Nicholas Lehman, Mauricio Di Fulvio, Nicholas McCray, Isabel Campos, Farnaz Tabatabaian, and Julian Gomez-Cambronero*
Cell Biology and Physiology, Wright State University School of Medicine, Dayton, Ohio
* Corresponding author; email: julian.cambronero{at}wright.edu.
We have investigated whether the signaling protein phospholipase D is implicated in leukocyte cell motility. Treating differentiated HL-60 cells with small interfering RNAs (siRNAs), in order to deplete endogenous expression of the PLD1 isoform, led to an abolishment of basal chemokinesis, that could not be rescued with chemoattractants ENA-78, FMLP and IL-8. Transient overexpression of PLD1 increased both chemokinesis and chemotaxis towards IL-8 and FMLP, but not towards ENA-78. Chemokinesis was not mediated by PLD1’ s enzymatic activity, but the chemotactic response was, as a lipase-inactive mutant (PLD1-K830R) negated all chemokine-induced potentiating actions; and since IL-8 and FMLP increased activity in vitro. Gene expression silencing of the other mammalian isoform, PLD2, also led to cell migration arrest, whereas ENA-78 selectively increased endogenous PLD2 activity and chemotaxis of HL-60s overexpressing a myc-pdDNA-PLD2 construct. Thus, PLD1 is differentially activated by CXCR-1, while CXCR-2 (and possibly CXCR-1) mediates PLD2 activation. Finally, immunofluorescence microscopy showed that both isoforms were associated with cell polarity and directionality concomitantly with adhesion and F-actin polymerization in response to IL-8. These data represent the first demonstration of the involvement of PLD and its enzymatic activities towards chemokines in the key physiological process of leukocyte migration.
