The long non-coding RNA GAS5 differentially regulates cell cycle arrest and apoptosis through activation of BRCA1 and p53 in human neuroblastoma

The long non-coding RNA GAS5 has been shown to modulate cancer proliferation in numerous human cancer systems and has been correlated with successful patient outcome. Our examination of GAS5 in neuroblastoma has revealed robust expression in both MYCN-amplified and non-amplified cell lines. Knockdown of GAS5 In vitro resulted in defects in cell proliferation, apoptosis, and induced cell cycle arrest. Further analysis of GAS5 clones revealed multiple novel splice variants, two of which inversely modulated with MYCN status. Complementation studies of the variants post-knockdown of GAS5 indicated alternate phenotypes, with one variant (FL) considerably enhancing cell proliferation by rescuing cell cycle arrest and the other (C2) driving apoptosis, suggesting a unique role for each in neuroblastoma cancer physiology. Global sequencing and ELISA arrays revealed that the loss of GAS5 induced p53, BRCA1, and GADD45A, which appeared to modulate cell cycle arrest in concert. Complementation with only the FL GAS5 clone could rescue cell cycle arrest, stabilizing HDM2, and leading to the loss of p53. Together, these data offer novel therapeutic targets in the form of lncRNA splice variants for separate challenges against cancer growth and cell death.

[1]  Erratum to: LncRNA-N1LR Enhances Neuroprotection Against Ischemic Stroke Probably by Inhibiting p53 Phosphorylation , 2017, Molecular Neurobiology.

[2]  Yang Xu,et al.  Regulation of p53 responses by post-translational modifications , 2003, Cell Death and Differentiation.

[3]  Tong Tong,et al.  Gadd45a contributes to p53 stabilization in response to DNA damage , 2003, Oncogene.

[4]  Zhuomin Wu,et al.  Erratum to: LncRNA-N1LR Enhances Neuroprotection Against Ischemic Stroke Probably by Inhibiting p53 Phosphorylation , 2017, Molecular Neurobiology.

[5]  F. Westermann,et al.  Functional Analysis of the p53 Pathway in Neuroblastoma Cells Using the Small-Molecule MDM2 Antagonist Nutlin-3 , 2011, Molecular Cancer Therapeutics.

[6]  P. Stadler,et al.  RNA Maps Reveal New RNA Classes and a Possible Function for Pervasive Transcription , 2007, Science.

[7]  Arsen O. Batagov,et al.  HOXD-AS1 is a novel lncRNA encoded in HOXD cluster and a marker of neuroblastoma progression revealed via integrative analysis of noncoding transcriptome , 2014, BMC Genomics.

[8]  P. May,et al.  Twenty years of p53 research: structural and functional aspects of the p53 protein , 1999, Oncogene.

[9]  S. Diederichs,et al.  Gutschner T , Diederichs S . The hallmarks of cancer : a long non-coding RNA point of view . RNA Biol 9 : 703-719 , 2012 .

[10]  Ming Sun,et al.  Decreased expression of long noncoding RNA GAS5 indicates a poor prognosis and promotes cell proliferation in gastric cancer , 2014, BMC Cancer.

[11]  Julie R. Park,et al.  Neuroblastoma: paradigm for precision medicine. , 2015, Pediatric clinics of North America.

[12]  A. Nakagawara,et al.  Transcriptional regulation of tumor suppressor p53 by cAMP‐responsive element‐binding protein/AMP‐activated protein kinase complex in response to glucose deprivation , 2009, Genes to cells : devoted to molecular & cellular mechanisms.

[13]  Q. Zhan.  Gadd45a, a p53- and BRCA1-regulated stress protein, in cellular response to DNA damage. , 2005, Mutation research.

[14]  C. Kanduri,et al.  Long noncoding RNAs and neuroblastoma , 2015, Oncotarget.

[15]  T. Ouchi,et al.  BRCA1 regulates p53-dependent gene expression. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[16]  Xuefei Shi,et al.  A critical role for the long non‐coding RNA GAS5 in proliferation and apoptosis in non‐small‐cell lung cancer , 2015, Molecular carcinogenesis.

[17]  M. Gatei,et al.  Role for ATM in DNA damage-induced phosphorylation of BRCA1. , 2000, Cancer research.

[18]  Björn Schumacher,et al.  The p53 network: cellular and systemic DNA damage responses in aging and cancer. , 2012, Trends in genetics : TIG.

[19]  Gwyn T. Williams,et al.  Regulation of apoptosis by long non-coding RNA GAS5 in breast cancer cells: implications for chemotherapy , 2014, Breast Cancer Research and Treatment.

[20]  Gaurav Kumar Pandey,et al.  The risk-associated long noncoding RNA NBAT-1 controls neuroblastoma progression by regulating cell proliferation and neuronal differentiation. , 2014, Cancer cell.

[21]  J. Sutcliffe,et al.  Primate brain-specific cytoplasmic transcript of the Alu repeat family , 1987, Molecular and cellular biology.

[22]  S. Jason,et al.  Neuroblastoma , 2018 .

[23]  William A Weiss,et al.  Neuroblastoma and MYCN. , 2013, Cold Spring Harbor perspectives in medicine.

[24]  R. Spizzo,et al.  Long non-coding RNAs and cancer: a new frontier of translational research? , 2012, Oncogene.

[25]  W. Lukiw,et al.  BC200 RNA in normal human neocortex, non-Alzheimer dementia (NAD), and senile dementia of the Alzheimer type (AD) , 1992, Neurochemical Research.

[26]  R. Arceci.  The International Neuroblastoma Risk Group (INRG) Classification System: An INRG Task Force Report , 2009 .

[27]  P. Ongusaha,et al.  BRCA1 shifts p53-mediated cellular outcomes towards irreversible growth arrest , 2003, Oncogene.

[28]  M. Mourtada-Maarabouni,et al.  Long non-coding RNA GAS5 regulates apoptosis in prostate cancer cell lines. , 2013, Biochimica et biophysica acta.

[29]  M. Kulesz-Martin,et al.  p53 protein at the hub of cellular DNA damage response pathways through sequence-specific and non-sequence-specific DNA binding. , 2001, Carcinogenesis.

[30]  M. Barton,et al.  Making sense of ubiquitin ligases that regulate p53 , 2010, Cancer biology & therapy.

[31]  F. Christians,et al.  Induction of GADD45 and JNK/SAPK-Dependent Apoptosis following Inducible Expression of BRCA1 , 1999, Cell.

[32]  S. Elledge,et al.  Requirement of ATM-dependent phosphorylation of brca1 in the DNA damage response to double-strand breaks. , 1999, Science.

[33]  Howard Y. Chang,et al.  Long noncoding RNAs and human disease. , 2011, Trends in cell biology.

[34]  C. Prives,et al.  p53: puzzle and paradigm. , 1996, Genes & development.

[35]  R. Stahel,et al.  GAS5 long non-coding RNA in malignant pleural mesothelioma , 2014, Molecular Cancer.

[36]  K. McHugh,et al.  Neuroblastoma in childhood: review and radiological findings , 2005, Cancer imaging : the official publication of the International Cancer Imaging Society.

[37]  Jun-Tao Jiang,et al.  Downregulation of GAS5 Promotes Bladder Cancer Cell Proliferation, Partly by Regulating CDK6 , 2013, PloS one.

[38]  Y Taya,et al.  A DNA damage signal is required for p53 to activate gadd45. , 2000, Cancer research.

[39]  R. Fåhraeus,et al.  p53-mediated control of gene expression via mRNA translation during Endoplasmic Reticulum stress , 2015, Cell cycle.

[40]  S. Kent,et al.  Enzymatic activity of a synthetic 99 residue protein corresponding to the putative HIV-1 protease , 1988, Cell.

[41]  D. Tweddle,et al.  p53, SKP2, and DKK3 as MYCN Target Genes and Their Potential Therapeutic Significance , 2012, Front. Oncol..

[42]  S. Aerts,et al.  p53 induces formation of NEAT1 lncRNA-containing paraspeckles that modulate replication stress response and chemosensitivity , 2016, Nature Medicine.

[43]  M. Mourtada-Maarabouni,et al.  GAS5, a non-protein-coding RNA, controls apoptosis and is downregulated in breast cancer , 2009, Oncogene.

[44]  D. Meek.  Tumour suppression by p53: a role for the DNA damage response? , 2009, Nature Reviews Cancer.

[45]  J. Mattick,et al.  Non-coding RNA. , 2006, Human molecular genetics.

[46]  Hui Tian,et al.  BRCA1 physically associates with p53 and stimulates its transcriptional activity , 1998, Oncogene.

[47]  Zhuohua Zhang,et al.  Detection of protein ubiquitination. , 2009, Journal of visualized experiments : JoVE.

[48]  G. Brodeur.  Neuroblastoma: biological insights into a clinical enigma , 2003, Nature Reviews Cancer.