Long non-coding (lnc)-RNAs are key signaling molecules controlling cell behavior. Of note, lncRNAs expression is largely deregulated in inherited and acquired disorders, such as cancers. This project focuses on the role of the lncRNA HOTAIR in the Epithelial-to-Mesenchymal transition (EMT), a transdifferentiation process that, allowing invasion of tumor cells in secondary sites, is required to metastasis. HOTAIR is overexpressed in several epithelial cancers and strongly correlated to invasion. Moreover, we have recently provided evidence that this lncRNA exerts a key role in the EMT triggered by the master factor Snail: this transcriptional repressor, sufficient to induce and maintain EMT, must bind to HOTAIR to recruit the Polycomb member EZH2 to specific targets (that will undergo EZH2-induced chromatin repressive modifications). Therefore, Snail-mediated repression of epithelial genes requires HOTAIR.
Building on this evidence, and as a continuation of previous researches, this project aims to: i) the design and the characterization of HOTAIR mutant molecules, potentially acting as dominant negative regulators of HOTAIR function; ii) the identification of HOTAIR partners and the study of their mechanism of action in the chromatin context. The proposed goals will clarify the regulatory circuitries in which HOTAIR participates; they will also define how the deregulation of this lncRNA impacts on cancer.
In gathering this body of evidence, we will acquire knowledge to be applied to successive development of biotechnological tools for innovative lncRNA-based therapeutic strategies in cancer.
Our hypothesis is buildt on our previous highly-cited description of HOTAIR role in EMT (Battistelli et al., Oncogene 2017, IF 2016 7,52; 54 Citations by Scopus, May 2019) and corroborated by further unpublished results of the group and preliminary bioinformatic analysis (in collaboration with Prof. Gian Gaetano Tartaglia, Dept. Biology and Biotechnology Charles Darwin, Sapienza, University of Rome) that identified by new approaches potential protein binding regions of murine and human HOTAIR RNAs. It is expected that the HOTAIR mutants, competing with the wild type HOTAIR for the Snail binding, interfere with the chromatin modifications causal to the repression of epithelial genes. This, in turn, should result in the loss of invasive properties and could interfere with the EMT execution. If the role of the HOTAIR mutants will be validated in human cells, these molecules could represent an innovative tool for gene therapy approaches counteracting epithelial tumor metastasis.
We also aim at the identification of genomic loci where HOTAIR is recruited in epithelial invasive cells and at the discovery of other protein interactors of this lncRNA in the chromatin context during EMT and tumor progression. This study will allow to extend our understanding on the role of HOTAIR in carcinogenesis, thus paving the way for the design of proper strategies counteracting its dominant role in EMT.
With respect to the feasibility, the proponent has a large experience in the characterization of molecular events underlying the EMT/MET dynamics of the epithelial cells, in physiological as well as pathological conditions, and this is highlighted by her specific and highly-cited publications on which large part of the proposed research builds. We also like to stress that all the techniques included in the experimental procedure are currently in use by the group. Furthermore, C C and CB set up all the innovative protocols used in previous studies for in vivo analyses of RNA/DNA/protein interactions in the chromatin context. We believe that the expertise of all partecipants in cellular and molecular biology will expedite successful advances in the proposed tasks.
Proteomic analysis will be performed in collaboration with Dr. C. Montaldo (Proteomic Unit, National Institute for Infectious Diseases L. Spallanzani, IRCCS, Rome, Italy).