Functional links between diacylglycerol and phosphatidylinositol signaling systems in human leukocyte-derived cell...

In leukocytes, diacylglycerol (DAG) regulates the small GTPase Ras through the agency of Ras guanyl releasing proteins (RasGRPs). Ras is thought to regulate phosphatidylinositol 3-kinase (PI3K). Therefore, DAG signaling is hypothesized to impact PI3K activity. The DAG analogue "pico" was used to activate RasGRPs in leukocyte-derived cell lines. PI3K signaling was monitored using antibodies that recognize the activated form of the PI3K-regulated protein kinase Akt. Diverse responses were documented. Some cell lines exhibit a DAG analogue-stimulated increase in phospho-Akt but this response proceeded even when Ras activation was blocked. In some Epstein-Barr virus-associated malignant cell lines and transformed B cells, high basal phospho-Akt was decreased by DAG analogue treatment. The pan-PKC inhibitor Bisindolymaleimide I blocked this effect. Basal phospho-Akt was also decreased by treatment with Go6976, an inhibitor of conventional protein kinase C and protein kinase D. While the proposed RasGRP-Ras-PI3K-Akt signaling axis may operate in some situations, our results indicate that alternative links between DAG targets such as protein kinases and the PI3K signaling system are more prominent.