AN INTEGRATED ROLE FOR CHROMATIN AND lncRNAs IN DEFINING CELL-SPECIFIC SPLICING PROGRAMS – EpiSplicing

The regulation of gene expression patterns, during development and in cell differentiation, plays an important role in cell commitment and the definition of cell identity. Many efforts have been dedicated to understand how cell-specific transcriptional programs are established and maintained. However in the past few years, another important regulatory layer has emerged, alternative splicing. Alternative splicing is a very general process essential for protein diversity that increases genome plasticity. A cell is thus no longer defined by the subset of genes that are transcribed, but as well, by how those genes are processed during alternative splicing, creating sophisticated developmental- and tissue-specific splicing programs essential for the correct function of the cell.
In this project, we aim to understand how cell-specific splicing programs are established and maintained, paying special attention to two innovative and newly identified types of splicing regulation: chromatin and long non-coding RNAs. Based on our previous publications and preliminary data, I hypothesize that lncRNAs and chromatin modifications play an important role in defining cell-specific splicing programs. For testing this hypothesis, we will combine state-of-the-art genome-wide approaches with molecular and cell biology techniques in a dynamic and physiologically relevant cellular model, the human epithelial-to-mesenchymal transition (EMT), involved in early development and cancer. In detail, we will integrate in time, during EMT, the changes in alternative splicing with changes in chromatin and lncRNAs along EMT-dependent splicing events. Our aim is to identify novel regulators of EMT-specific splicing that will be functionally characterized by adapting the innovative CRISPR/Cas9 system, and other state-of-the-art techniques, to mechanistically understand the role of chromatin and lncRNAs in the regulation of alternative splicing during EMT. Finally the candidates that can best revert EMT-dependent changes in splicing will be further tested for their capacity to impair EMT progression itself in vitro and in vivo in mouse. Results from this research will provide fundamental understanding of the mechanisms controlling cell-specific splicing and the integrative role of chromatin and lncRNAs in gene expression regulation in an inducible and biologically relevant system, the EMT. This project is therefore perfectly fitting the "Societal Challenge: Life, Health and Well-being", and in particular the "Axe 5: Genetics, Genomics and Epigenetics", by integrating multidisciplinary approaches, from genome-wide transcriptomics and epigenomics to innovative molecular and cancer biology techniques, to depict the dynamic interplay between the multiple levels of gene regulation during the establishment and maintenance of a new cell-specific program involved in tumour progression. Moreover, these studies aim to discover unsuspected new mechanisms of cell-specific splicing regulation, using a physiologically relevant cellular model system, the EMT. In the long term, results from this project will bring new perspectives into the development of more gene-specific and accurate therapeutic strategies based on the use of non-coding RNAs to revert disease-specific splicing programs and improve cancer prognosis. As an evidence of the potential economical and societal impact of this project, there are already American Spin offs, such as RaNa Therapeutics and Isis Pharmaceuticals, investing in the use of lncRNAs and antisense DNA to treat splicing-specific diseases, such as Muscular Dystrophy.