LPS induces phosphorylation of the transcription factors ATF-2 and CREB in a p38SAPK-dependent manner.

Peripheral blood monocytes cultured with GM-CSF and IL-4 for 7 days (immature MoDCs) were exposed to LPS (100 ng/mL), and after variable lengths of time, samples were removed, separated by SDS/PAGE, and then probed with phosphorylation-specific antibodies to the transcription factors CREB and ATF-2. In some cases, the day-7 MoDCs were preincubated with the p38SAPK inhibitor SB203580 for 30 minutes before exposure to LPS. Because SB203580 was dissolved in DMSO, MoDCs that had been incubated with DMSO alone were used as controls (d). The Western blots show that phosphorylation (and hence activation) of the transcription factors ATF-2 and CREB is induced by the addition of LPS within 30 minutes. This appears to be mediated via the p38SAPK pathway because blocking this prevents the LPS-induced phosphorylation of these transcription factors. The faint bands that appear below the phospho-CREB bands in the middle panel are due to cross-reactivity of the anti-phospho–ATF-2 antibody with ATF-1, which runs in this position. The lowest panel shows the blot probed for total ATF-2 and CREB to demonstrate loading of samples. Similar results were obtained in 3 separate experiments.