Modification of cysteine residue in p65 subunit of nuclear factor-kappaB (NF-kappaB) by picroliv suppresses NF-kappaB-regulated gene products and potentiates apoptosis.
Picroliv, an iridoid glycoside derived from the plant Picrorhiza kurroa, is used traditionally to treat fever, asthma, hepatitis, and other inflammatory conditions. However, the exact mechanism of its therapeutic action is still unknown. Because nuclear factor-kappaB (NF-kappaB) activation plays a major role in inflammation and carcinogenesis, we postulated that picroliv must interfere with this pathway by inhibiting the activation of NF-kappaB-mediated signal cascade. Electrophoretic mobility shift assay showed that pretreatment with picroliv abrogated tumor necrosis factor (TNF)-induced activation of NF-kappaB. The glycoside also inhibited NF-kappaB activated by carcinogenic and inflammatory agents, such as cigarette smoke condensate, phorbol 12-myristate 13-acetate, okadaic acid, hydrogen peroxide, lipopolysaccharide, and epidermal growth factor. When examined for the mechanism of action, we found that picroliv inhibited activation of IkappaBalpha kinase, leading to inhibition of phosphorylation and degradation of IkappaBalpha. It also inhibited phosphorylation and nuclear translocation of p65. Further studies revealed that picroliv directly inhibits the binding of p65 to DNA, which was reversed by the treatment with reducing agents, suggesting a role for a cysteine residue in interaction with picroliv. Mutation of Cys(38) in p65 to serine abolished this effect of picroliv. NF-kappaB inhibition by picroliv leads to suppression of NF-kappaB-regulated proteins, including those linked with cell survival (inhibitor of apoptosis protein 1, Bcl-2, Bcl-xL, survivin, and TNF receptor-associated factor 2), proliferation (cyclin D1 and cyclooxygenase-2), angiogenesis (vascular endothelial growth factor), and invasion (intercellular adhesion molecule-1 and matrix metalloproteinase-9). Suppression of these proteins enhanced apoptosis induced by TNF. Overall, our results show that picroliv inhibits the NF-kappaB activation pathway, which may explain its anti-inflammatory and anticarcinogenic effects.
Anand
Preetha
P
Department of Experimental Therapeutics, Cytokine Research Laboratory, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Kunnumakkara
Ajaikumar B
AB
Harikumar
Kuzhuvelil B
KB
Ahn
Kwang Seok
KS
Badmaev
Vladimir
V
Aggarwal
Bharat B
BB
D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002934:Cinnamates / Q000494:pharmacology*; D003545:Cysteine / Q000737:chemistry*; D004273:DNA, Neoplasm / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D024202:Electrophoretic Mobility Shift Assay; D006027:Glycosides / Q000494:pharmacology*; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D016328:NF-kappa B / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D021381:Protein Transport; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology; D014641:Vanillic Acid / Q000494:pharmacology*
D002934:Cinnamates; D004273:DNA, Neoplasm; D006027:Glycosides; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; C057463:kutkin; D014641:Vanillic Acid; D003545:Cysteine