Phosphatidic acid mobilized by phospholipase D is involved in the phorbol 12-myristate 13-acetate-induced G2 delay of A431 cells.

This study was aimed at gaining an understanding of metabolic events responsible for the inhibition of cells in G2 phase, a known physiological restriction site in the cell cycle of multicellular organisms. In an earlier study, phosphatidic acid was proposed as an inhibitory mediator in the epidermal growth factor (EGF)-induced inhibition of A431 cells in G2 phase via the phospholipase C pathway [Kaszkin, Richards and Kinzel (1992) Cancer Res. 52, 5627-5634]. We show here that the phorbol ester phorbol 12-myristate 13-acetate (PMA) induces a reversible inhibition of the G2/M transition in A431 cells under conditions of phospholipase D-catalysed phosphatidic acid formation. Such PMA-induced inhibition in G2 phase is largely attenuated in the presence of 1-propanol (but not of 2-propanol). In this case the amount of phosphatidic acid is reduced to almost control levels, and instead phosphatidylpropanol is formed. In the case of EGF-induced activation of a phospholipase D the amount of phosphatidic acid is only slightly decreased in the presence of a primary alcohol. Under these conditions the EGF-induced G2 delay was not affected. The correlation between the formation of phosphatidic acid and the G2 delay induced by PMA, as well as by an exogenous bacterial phospholipase D (from Streptomyces chromofuscus), could be supported by using synchronized cells in order to increase the population of cells in G2 phase. This study indicates that the formation of substantial amounts of phosphatidic acid immediately before entry into mitosis seems to be important for establishing a delay in the cell cycle at the G2/M border by exogenous ligands.