Identification and analysis of non-coding loci critical for cell viability and chromatin organization

Project Summary: The effort to identify functional non-coding sequences has largely focused on the regulatory elements such as enhancers and non-coding RNAs. However, while ~70% of the human genome is epigenetically quiescent, i.e. not marked by any epigenetic modification indicating functional inactivity, the majority of somatic mutations associated with diseases occur in the non-coding regions. An emerging question is why and how somatic mutations in epigenetically quiescent loci would affect cellular functions and cause disease formation. A possible mechanism is that the mutations in these loci may alter the neighbor chromatin organization and such a change is propagated through the 3D genome to generate a profound impact on phenotype. Although this hypothesis is tempting, such non-coding loci have not been widely identified, characterized and analyzed in the human genome. In the proposed work, we aim to integrate computational prediction and CRISPR library screen to systematically uncover non-coding loci that do not host any functional element and are not even marked by histone modification, protein binding or open chromatin but are important for cell survival (Specific Aim 1 and 2). We will investigate the mechanisms of how the deletion of these loci lead to cell dysfunction using Hi-C, single cell RNA-seq and single ATAC-seq analyses (Specific Aim 3). Once completed, the proposed work will provide a new aspect of understanding the functions of epigenetically quiescent non-coding loci and facilitate developing genome-editing based therapeutics targeting these loci.