Background: Platelet-derived growth factor (PDGF) triggers cytoskeletal rearrangements and chemotaxis within minutes. These events are at least in part due to the activation of phosphoinositide (PI) 3-kinase; there is good temporal correlation between these events and the accumulation of 3-phosphorylated products of the kinase. Prolonged and continuous PDGF exposure results in S-phase entry many hours after the initial burst of activity. Although early signals appear responsible for the early responses, they may not fully account for later responses, such as cell-cycle progression.
Results: We assessed when PI 3-kinase products accumulate in PDGF-stimulated cells. In addition to the previously identified early accumulation of products, we detected a second, prolonged wave of accumulation 3-7 hours after stimulation. To determine the relative contribution of each phase to PDGF-dependent DNA synthesis, we first developed an assay in which synthetic 3-phosphorylated lipids were used to rescue DNA synthesis in cells expressing a PDGF-receptor mutant. The lipids rescued DNA synthesis only when added 2-6 hours after PDGF. In addition, PI 3-kinase inhibitors failed to block PDGF-dependent DNA synthesis if added during the first wave of PI 3-kinase activity, but adding them later, in G1 phase, prevented PDGF-dependent cell-cycle progression.
Conclusions: PDGF induces distinct waves of PI 3-kinase activity. The second wave is required for PDGF-dependent DNA synthesis, whereas the initial wave is not. One of the ways in which cells use PI 3-kinase to mediate distinct cellular responses seems to be by regulating when its products accumulate.