Suppression of RelA/p65 transactivation activity by a lignoid manassantin isolated from Saururus chinensis

In our search for NF-kappaB inhibitors from natural resources, we have previously identified two structurally related dilignans, manassantin A and B as specific inhibitors of NF-kappaB activation from Saururus chinensis. However, their molecular mechanism of action remains unclear. We here demonstrate that manassantins A and B are potent inhibitors of NF-kappaB activation by the suppression of transciptional activity of RelA/p65 subunit of NF-kappaB. These compounds significantly inhibited the induced expression of NF-kappaB reporter gene by LPS or TNF-alpha in a dose-dependent manner. However, these compounds did not prevent the DNA-binding activity of NF-kappaB assessed by electrophoretic mobility shift assay as well as the induced-degradation of IkappaB-alpha protein by LPS or TNF-alpha. Further analysis revealed that manassantins A and B dose-dependently suppressed not only the induced NF-kappaB activation by overexpression of RelA/p65, but also transactivation activity of RelA/p65. Furthermore, treatment of cells with these compounds prevented the TNF-alpha-induced expression of anti-apoptotic NF-kappaB target genes Bfl-1/A1, a prosurvival Bcl-2 homologue, and resulted in sensitizing HT-1080 cells to TNF-alpha-induced cell death. Similarly, these compounds also suppressed the LPS-induced inducible nitric oxide synthase expression and nitric oxide production. Taken together, manassantins A and B could be valuable candidate for the intervention of NF-kappaB-dependent pathological condition such as inflammation and cancer.