PHOSPHATIDIC-ACID MOBILIZED BY PHOSPHOLIPASE-D IS INVOLVED IN THE PHORBOL 12-MYRISTATE 13-ACETATE-INDUCED G(2) DELAY OF A431 CELLS

This study was aimed at gaining an understanding of metabolic events r esponsible for the inhibition of cells in G(2) phase, a known physiolo gical restriction site in the cell cycle of multicellular organisms. I n an earlier study, phosphatidic acid was proposed as an inhibitory me diator in the epidermal growth factor (EGF)-induced inhibition of A431 cells in G(2) phase via the phospholipase C pathway [Kaszkin, Richard s and Kinzel (1992) Cancer Res. 52, 5627-5634]. We show here that the phorbol ester phorbol 12-myristate 13-acetate (PMA) induces a reversib le inhibition of the G(2)/M transition in A431 cells under conditions of phospholipase D-catalysed phosphatidic acid formation. Such PMA-ind uced inhibition in G(2) phase is largely attenuated in the presence of 1-propanol (but not of 2-propanol). In this case the amount of phosph atidic acid is reduced to almost control levels, and instead phosphati dylpropanol is formed. In the case of EGF-induced activation of a phos pholipase D the amount of phosphatidic acid is only slightly decreased in the presence of a primary alcohol. Under these conditions the EGF- induced G(2) delay was not affected. The correlation between the forma tion of phosphatidic acid and the G(2) delay induced by PMA, as well a s by an exogenous bacterial phospholipase D (from Streptomyces chromo- fuscus), could be supported by using synchronized cells in order to in crease the population of cells in G(2) phase. This study indicates tha t the formation of substantial amounts of phosphatidic acid immediatel y before entry into mitosis seems to be important for establishing a d elay in the cell cycle at the G(2)/M border by exogenous ligands.