Mitogenic phospholipase D activity is restricted to caveolin-enriched membrane microdomains

Phospholipase D (PLD) activity is elevated in response to the oncogenic sti mulus of several signaling oncogenes. PLD activity is also elevated in resp onse to peptide growth factors, indicating that PLD likely plays an importa nt role in mitogenic signaling. Many proteins that mediate mitogenic signal ing are localized in caveolin-enriched membrane microdomains (CEMMs). We re port here that the elevated PLD activity in NIH 3T3 cells transformed by ac tivated oncogenic forms of Src, Res, and Raf is largely restricted to the C EMMs. Likewise, the PLD activity stimulated by epidermal growth factor is a lso restricted to the CEMMs. Although both PLD1 and PLD2 were found in CEMM s, neither was particularly enriched in the CEMMs of the transformed relati ve to the parental cells, indicating that it is the specific activity of PL D that is increased in the CEMMs. An apparent PLD substrate specificity in transformed cells for phosphatidylcholine lacking arachidonate acyl groups is also explained by the localization of activity in the CEMMs where [H-3]a rachidonate-labeled PC was excluded. These data indicate that mitogenic sig nals through PLD are initiated in CEMMs where many signaling molecules colo calize. (C) 2000 Academic Press.