Long non-coding RNA MEG3 inhibits NSCLC cells proliferation and induces apoptosis by affecting p53 expression

BackgroundLong non-coding RNAs play an important role in tumorigenesis, hence, identification of cancer-associated lncRNAs and investigation of their biological functions and molecular mechanisms are important for understanding the development and progression of cancer. Recently, the downregulation of lncRNA MEG3 has been observed in various human cancers. However, its role in non-small cell lung cancer (NSCLC) is unknown. The aim of this study was to examine the expression pattern of MEG3 in NSCLC and to evaluate its biological role and clinical significance in tumor progression.MethodsExpression of MEG3 was analyzed in 44 NSCLC tissues and 7 NSCLC cell lines by qRT-PCR. Over-expression approaches were used to investigate the biological functions of MEG3 in NSCLC cells. Bisulfite sequencing was used to investigate DNA methylation on MEG3 expression. The effect of MEG3 on proliferation was evaluated by MTT and colony formation assays, and cell apoptosis was evaluated by Hoechst staining and Flow-cytometric analysis. NSCLC cells transfected with pCDNA-MEG3 were injection into nude mice to study the effect of MEG3 on tumorigenesis in vivo . Protein levels of MEG3 targets were determined by western blot analysis. Differences between groups were tested for significance using Student’s t-test (two-tailed).ResultsMEG3 expression was decreased in non-small cell lung cancer (NSCLC) tumor tissues compared with normal tissues, and associated with advanced pathologic stage, and tumor size. Moreover, patients with lower levels of MEG3 expression had a relatively poor prognosis. Overexpression of MEG3 decreased NSCLC cells proliferation and induced apoptosis in vitro and impeded tumorigenesis in vivo. MDM2 and p53 protein levels were affected by MEG3 over-expression in vitro.ConclusionsOur findings indicate that MEG3 is significantly down-regulated in NSCLC tissues that could be affected by DNA methylation, and regulates NSCLC cell proliferation and apoptosis, partially via the activition of p53. Thus, MEG3 may represent a new marker of poor prognosis and is a potential therapeutic target for NSCLC intervention.

[1]  Yunli Zhou,et al.  MEG3 noncoding RNA: a tumor suppressor. , 2012, Journal of molecular endocrinology.

[2]  J. Rinn,et al.  Large intergenic non-coding RNA-RoR modulates reprogramming of human induced pluripotent stem cells , 2010, Nature Genetics.

[3]  Chen Yang,et al.  Long non-coding RNA UCA1a(CUDR) promotes proliferation and tumorigenesis of bladder cancer. , 2012, International journal of oncology.

[4]  Yong Huang,et al.  Regulatory long non-coding RNA and its functions , 2012, Journal of Physiology and Biochemistry.

[5]  J. Rinn,et al.  Many human large intergenic noncoding RNAs associate with chromatin-modifying complexes and affect gene expression , 2009, Proceedings of the National Academy of Sciences.

[6]  Jing Zhao,et al.  Activation of p53 by MEG3 Non-coding RNA* , 2007, Journal of Biological Chemistry.

[7]  Jianfeng Xu,et al.  Human polymorphisms at long non-coding RNAs (lncRNAs) and association with prostate cancer risk. , 2011, Carcinogenesis.

[8]  A. Jemal,et al.  Cancer Statistics, 2010 , 2010, CA: a cancer journal for clinicians.

[9]  Ming Sun,et al.  MicroRNA-196a promotes non-small cell lung cancer cell proliferation and invasion through targeting HOXA5 , 2012, BMC Cancer.

[10]  Yiran Huang,et al.  Upregulated MALAT-1 contributes to bladder cancer cell migration by inducing epithelial-to-mesenchymal transition. , 2012, Molecular bioSystems.

[11]  Jiayi Wang,et al.  CREB up-regulates long non-coding RNA, HULC expression through interaction with microRNA-372 in liver cancer , 2010, Nucleic acids research.

[12]  Michael F. Lin,et al.  Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals , 2009, Nature.

[13]  E. Hatzimichael,et al.  MEG3 imprinted gene contribution in tumorigenesis , 2011, International journal of cancer.

[14]  Pengjun Wang,et al.  Overexpression of the long non‐coding RNA MEG3 impairs in vitro glioma cell proliferation , 2012, Journal of cellular biochemistry.

[15]  C. Ponting,et al.  Evolution and Functions of Long Noncoding RNAs , 2009, Cell.

[16]  David G. Knowles,et al.  The GENCODE v7 catalog of human long noncoding RNAs: Analysis of their gene structure, evolution, and expression , 2012, Genome research.

[17]  Shuhan Sun,et al.  Long noncoding RNA associated with microvascular invasion in hepatocellular carcinoma promotes angiogenesis and serves as a predictor for hepatocellular carcinoma patients' poor recurrence‐free survival after hepatectomy , 2012, Hepatology.

[18]  J. Rinn,et al.  A Large Intergenic Noncoding RNA Induced by p53 Mediates Global Gene Repression in the p53 Response , 2010, Cell.

[19]  William Stafford Noble,et al.  Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project , 2007, Nature.

[20]  Leila Mohammadi,et al.  BMC Cancer , 2001 .

[21]  Hermann Brenner,et al.  Recent cancer survival in Europe: a 2000-02 period analysis of EUROCARE-4 data. , 2007, The Lancet. Oncology.

[22]  Shuhan Sun,et al.  Long noncoding RNA high expression in hepatocellular carcinoma facilitates tumor growth through enhancer of zeste homolog 2 in humans , 2011, Hepatology.

[23]  C. Prives,et al.  Blinded by the Light: The Growing Complexity of p53 , 2009, Cell.

[24]  Q. Li,et al.  Large Intervening Non-Coding RNA HOTAIR is Associated with Hepatocellular Carcinoma Progression , 2011, The Journal of international medical research.

[25]  A. Wierzbicki.  The role of long non-coding RNA in transcriptional gene silencing. , 2012, Current opinion in plant biology.

[26]  A. Ferguson-Smith,et al.  Epigenetic alteration at the DLK1-GTL2 imprinted domain in human neoplasia: analysis of neuroblastoma, phaeochromocytoma and Wilms' tumour , 2005, British Journal of Cancer.

[27]  S. Safe,et al.  HOTAIR IS A NEGATIVE PROGNOSTIC FACTOR AND EXHIBITS PRO-ONCOGENIC ACTIVITY IN PANCREATIC CANCER , 2012, Oncogene.

[28]  D. Spector,et al.  The noncoding RNA MALAT1 is a critical regulator of the metastasis phenotype of lung cancer cells. , 2013, Cancer research.

[29]  D. S. Gross,et al.  Chromatin , 2015, Current Biology.

[30]  I. Cuthill,et al.  Animal Research: Reporting In Vivo Experiments: The ARRIVE Guidelines , 2010, British journal of pharmacology.

[31]  J. Mattick,et al.  The melanoma-upregulated long noncoding RNA SPRY4-IT1 modulates apoptosis and invasion. , 2011, Cancer research.

[32]  Chaojun Li,et al.  Knockdown of c-Met by adenovirus-delivered small interfering RNA inhibits hepatocellular carcinoma growth in vitro and in vivo , 2005, Molecular Cancer Therapeutics.

[33]  Sergio Verjovski-Almeida,et al.  Long intronic noncoding RNA transcription: expression noise or expression choice? , 2009, Genomics.

[34]  D. Spector,et al.  Long noncoding RNAs: functional surprises from the RNA world. , 2009, Genes & development.

[35]  Howard Y. Chang,et al.  Long noncoding RNA HOTAIR reprograms chromatin state to promote cancer metastasis , 2010, Nature.

[36]  Howard Y. Chang,et al.  RNA templating the epigenome , 2011, Epigenetics.

[37]  C. Croce,et al.  microRNA-29 can regulate expression of the long non-coding RNA gene MEG3 in hepatocellular cancer , 2011, Oncogene.

[38]  Chuanliang Xu,et al.  The prostate cancer-up-regulated long noncoding RNA PlncRNA-1 modulates apoptosis and proliferation through reciprocal regulation of androgen receptor. , 2013, Urologic oncology.