Prodigiosin down-regulates SKP2 to induce p27KIP1 stabilization and antiproliferation in human lung adenocarcinoma cells

BACKGROUND AND PURPOSE High levels of SKP2 are a poor prognostic factor in multiple human cancers and mostly correlate with low p27KIP1 levels. Prodigiosin is a bacterial tripyrrole pigment with strong pro-apoptotic activity. Induction of cell cycle blockade underlies one of its anticancer actions but the mechanisms involved are unclear. The aim of this study was to explore the role of the SKP2p27KIP1 axis in prodigiosin's cytostatic effect on human lung adenocarcinoma cells. EXPERIMENTAL APPROACH Prodigiosin's effects on cell cycle progression and long-term cell proliferation of human lung adenocarcinoma cells were characterized by flow cytometry and colony formation assay, respectively. Real-time RT-PCR and promoter activity analyses were performed for assessing transcriptional control, while cycloheximide chase analysis evaluated protein stability. Immunoblotting was employed for mechanistic study. KEY RESULTS Prodigiosin increased p27KIP1 expression mainly by stabilizing p27KIP1 through transcriptional repression of SKP2. Importantly, SKP2 overexpression or p27KIP1 depletion restored the colony forming capacity of prodigiosin-treated cells. Furthermore, prodigiosin induced PKB dephosphorylation, leading to PKB inhibition as revealed by decreased serine 9 phosphorylation of GSK-3 beta. Constitutive PKB activation reduced prodigiosin-induced SKP2 repression. Prodigiosin also down-regulated E2F1 (mediates PI3K/PKB-induced SKP2 transcription), but E2F1 overexpression failed to restore SKP2 expression in prodigiosin-treated cells. CONCLUSIONS AND IMPLICATIONS Transcriptional repression of SKP2 and the consequent accumulation of p27KIP1 are essential for prodigiosin's antiproliferative action. Mechanistically, prodigiosin induces PKB inhibition to down-regulate SKP2 in a GSK-3 beta- and E2F1-independent manner. Our findings further implicate the potential for developing prodigiosin as a novel class of SKP2-targeting anticancer agent.