We have explored the levels of phosphatidylinositol-3 kinase protein during culture of rat 3Y1 cells. Confluent cell cultures exhibited a higher level of phosphatidylinositol-3 kinase compared with that of growing cells. After replating of the cells on fresh dishes, the level of phosphatidylinositol-3 kinase returned to that of growing cells within 24h. This density-dependent regulation of the phosphatidylinositol-3 kinase level was not lined to cell growth, because growth arrest by serum starvation did not cause elevation of the phosphatidylinositol-3 kinase level. Northern blotting analysis revealed that this regulation was based on the transcriptional level. After cell growth was arrested by contact inhibition, elevation of the level of phosphatidylinositol-3,4,5-trisphosphate was detected suggesting that phosphatidylinositol-3 kinase was activated in these cells. These effects were not seen in src-transformed 3Y1 cells, suggesting that this regulation was lost in transformed cells.