The lncRNA H19 positively affects the tumorigenic properties of glioblastoma cells and contributes to NKD1 repression through the recruitment of EZH2 on its promoter

// Barbara Fazi 1 , Sabrina Garbo 1 , Nicola Toschi 1, 2, 3 , Annunziato Mangiola 4 , Malinska Lombari 1 , Daria Sicari 1, 7 , Cecilia Battistelli 5 , Silvia Galardi 1 , Alessandro Michienzi 1 , Gianluca Trevisi 4 , Rona Harari-Steinfeld 6 , Carla Cicchini 5 and Silvia Anna Ciafre 1 1 Department of Biomedicine and Prevention, University of Rome “Tor Vergata”, Rome, Italy 2 Department of Radiology, Athinoula A. Martinos Center for Biomedical Imaging, Boston, MA, USA 3 Harvard Medical School, Boston, MA, USA 4 Department Head and Neck, Institute of Neurosurgery, Catholic University of Sacred Heart, Rome, Italy 5 Department of Cellular Biotechnologies and Haematology, Sezione di Genetica Molecolare, Istituto Pasteur Italia-Fondazione Cenci Bolognetti, Sapienza University of Rome, Rome, Italy 6 Goldyne Savad Institute of Gene Therapy, Hadassah University Hospital, Hebrew University, Jerusalem 7 Present address: Laboratorio Nazionale CIB (LNCIB), AREA Science Park, Trieste, Italy Correspondence to: Silvia Anna Ciafre, email: ciafre@uniroma2.it Keywords: glioblastoma; lncRNA; H19; NKD1; EZH2 Received: July 21, 2017 Accepted: February 10, 2018 Epub: February 14, 2018 Published: March 20, 2018 ABSTRACT The still largely obscure molecular events in the glioblastoma oncogenesis, a primary brain tumor characterized by an inevitably dismal prognosis, impel for investigation. The importance of Long noncoding RNAs as regulators of gene expression has recently become evident. Among them, H19 has a recognized oncogenic role in several types of human tumors and was shown to correlate to some oncogenic aspects of glioblastoma cells. Here we, hypothesyze that in glioblastoma H19 exerts its function through the interaction with the catalytic subunit of the PRC2 complex, EZH2. By employing a factor analysis on a SAGE dataset of 12 glioblastoma samples, we show that H19 expression in glioblastoma tissues correlates with that of several genes involved in glioblastoma growth and progression. H19 knock-down reduces viability, migration and invasiveness of two distinct human glioblastoma cell lines. Most importantly, we provide a mechanistic perspective about the role of H19 in glioblastoma cells, by showing that its expression is inversely linked to that of NKD1, a negative regulator of Wnt pathway, suggesting that H19 might regulate NKD1 transcription via EZH2-induced H3K27 trimethylation of its promoter. Indeed, we showed that H19 binds EZH2 in glioblastoma cells, and that EZH2 binding to NKD1 and other promoters is impaired by H19 silencing. In this work we describe H19 as part of an epigenetic modulation program executed by EZH2, that results in the repression of Nkd1. We believe that our results can provide a new piece to the complex puzzle of H19 function in glioblastoma.