Tanshinone IIA triggers p53 responses and apoptosis by RNA polymerase II upon DNA minor groove binding.
Our previous work has shown that tanshinone IIA (Tan IIA) is a DNA minor groove binder instead of an intercalator as previously thought. In this study, we have further demonstrated that the molecular antitumor pharmacology of Tan IIA is dependent on its groove-binding capability. First, we investigated the structure damage to duplex DNA upon Tan IIA binding using circular dichroism spectra. Subsequently, we performed western blot, flow cytometry analysis, chromatin immunoprecipitation, and quantitative real-time PCR to illustrate the RNAPII degradation, phosphorylation, and distribution along the transcribed gene in H22 cells exposed to Tan IIA. In addition, p53 activation and apoptosis induction in both cultured H22 cells and in mice bearing the ascitic-type H22 were measured following Tan IIA treatment. It was revealed that Tan IIA decreases the level of RNAPII by altering DNA structure. At the low dose range (0.2-4 microM) of Tan IIA exposure, the DNA structure damage results in the inhibition of RNAPII binding to DNA and the initiation of RNAPII phosphorylation, while higher concentrations of Tan IIA (4-20 microM) cause complete phosphorylation and degradation of RNAPII followed by p53 activation and apoptosis. A similar apoptosis induction by RNAPII was observed in animals. Apoptosis of tumor cells from ascitic fluid was not detected until RNAPII levels were downregulated by Tan IIA, which requires 40 mg/kg body weight of Tan IIA. It was concluded that DNA-conformational-damage-dependent RNAPII response upon groove binding is the molecular basis of the antitumor property of Tan IIA, in vivo and in vitro.
Zhang
Zhichao
Z
State Key Laboratory of Fine Chemicals, Dalian University of Technology, Dalian 116012, PR China. zczhang@dlut.edu.cn
Gao
Jin
J
Wang
Yuanyuan
Y
Song
Ting
T
Zhang
Jing
J
Wu
Guiye
G
Zhang
Tiantai
T
Du
Guanhua
G
D045784:Abietanes; D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000737:chemistry / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001202:Ascitic Fluid / Q000166:cytology / Q000378:metabolism; D001665:Binding Sites; D015153:Blotting, Western; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D020287:In Situ Nick-End Labeling; D008114:Liver Neoplasms, Experimental / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D009368:Neoplasm Transplantation; D009690:Nucleic Acid Conformation; D010616:Phenanthrenes / Q000737:chemistry / Q000494:pharmacology*; D010766:Phosphorylation; D012319:RNA Polymerase II / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D045784:Abietanes; D000972:Antineoplastic Agents, Phytogenic; D010616:Phenanthrenes; D016159:Tumor Suppressor Protein p53; C021751:tanshinone; D004247:DNA; D012319:RNA Polymerase II