title: Quantitative nuclear proteomics identifies that miR-137-mediated EZH2 reduction regulates resveratrol-induced apoptosis of neuroblastoma cells.
abstract: Neuroblastoma is the most common pediatric extracranial solid tumor with a broad spectrum of clinical behavior and poor prognosis. Despite intensive multimodal therapy, ongoing clinical trials, and basic science investigations, neuroblastoma remains a complex medical challenge with a long-term survival rate less than 40%. In our study, we found that resveratrol (3, 5, 4'-trihydroxystilbene, RSV), a naturally occurring phytoalexin, possesses an anticancer activity through blocking cell growth and inducing apoptosis in neuroblastoma cell line Neuro-2a (N-2a) cells. Using stable isotope labeling with amino acids in cell culture (SILAC) and quantitative proteomic analysis, we found that 395 proteins were up-regulated and 302 proteins were down-regulated in the nucleus of N-2a cells treated with RSV. Among these, the polycomb protein histone methyltransferase EZH2 was reduced significantly, which is aberrantly overexpressed in neuroblastoma and crucial to maintain the malignant phenotype of neuroblastoma by epigenetic repression of multiple tumor suppressor genes. EZH2 reduction further led to decreased H3K27me3 level and reactivation of neuroblastoma tumor suppressor genes CLU and NGFR. Disruption EZH2 expression by RNA interference-mediated knockdown or pharmacologic inhibition with DZNep triggered cellular apoptosis in N-2a cells. We found that the up-regulation of miR-137 level was responsible for reduced EZH2 level in tumor suppression induced by RSV. Inhibition of miR-137 expression rescued the cellular apoptosis phenotypes, EZH2 reduction, and CLU and NGFR reactivation, associated with RSV treatment. Taken together, our findings present for the first time, an epigenetic mechanism involving miR-137-mediated EZH2 repression in RSV-induced apoptosis and tumor suppression of neuroblastoma, which would provide a key potential therapeutic target in neuroblastoma treatment.
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China; ¶Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871, China;
authors.forename: Xiaoqing
authors.initials: X
authors.lastname: Ren
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Xue
authors.initials: X
authors.lastname: Bai
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Xuefei
authors.initials: X
authors.lastname: Zhang
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Zheyi
authors.initials: Z
authors.lastname: Li
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Lingfang
authors.initials: L
authors.lastname: Tang
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China; §Institute of System Biomedicine, School of Basic Medical Sciences, Peking University, Beijing 100191, China;
authors.forename: Xuyang
authors.initials: X
authors.lastname: Zhao
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Zeyang
authors.initials: Z
authors.lastname: Li
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China;
authors.forename: Yanfei
authors.initials: Y
authors.lastname: Ren
authors.affiliation: ¶Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871, China; ‖Department of Oral and Maxillofacial Surgery, Laboratory of Interdisciplinary Studies, School of Stomatology, Peking University, Beijing 100081, China;
authors.forename: Shicheng
authors.initials: S
authors.lastname: Wei
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China; wangqingsong@pku.edu.cn jijg@pku.edu.cn lcmicro@yahoo.com.
authors.forename: Qingsong
authors.initials: Q
authors.lastname: Wang
authors.affiliation: ‡‡Laboratory of Genome Stability, West China Second University Hospital, Sichuan University, Chengdu 610041, China wangqingsong@pku.edu.cn jijg@pku.edu.cn lcmicro@yahoo.com.
authors.forename: Cong
authors.initials: C
authors.lastname: Liu
authors.affiliation: From the ‡State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China; §Institute of System Biomedicine, School of Basic Medical Sciences, Peking University, Beijing 100191, China; wangqingsong@pku.edu.cn jijg@pku.edu.cn lcmicro@yahoo.com.
authors.forename: Jianguo
authors.initials: J
authors.lastname: Ji
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D019295:Computational Biology; D000071221:Enhancer of Zeste Homolog 2 Protein; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D007553:Isotope Labeling; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism*; D009447:Neuroblastoma / Q000378:metabolism* / Q000473:pathology; D009687:Nuclear Proteins / Q000378:metabolism*; D063151:Polycomb Repressive Complex 2 / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D020543:Proteome / Q000378:metabolism; D040901:Proteomics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D000077185:Resveratrol; D013267:Stilbenes / Q000494:pharmacology*; D013997:Time Factors
chemical_list: C551799:MIRN137 microRNA, mouse; D035683:MicroRNAs; D009687:Nuclear Proteins; D020543:Proteome; D012333:RNA, Messenger; D013267:Stilbenes; D000071221:Enhancer of Zeste Homolog 2 Protein; C498410:Ezh2 protein, mouse; D063151:Polycomb Repressive Complex 2; D000077185:Resveratrol