Very long intergenic non-coding (vlinc) RNAs directly regulate multiple genes in cis and trans

Majority of the human genome is transcribed in a form of long non-coding (lnc) RNAs. While these transcripts attracted considerable interest, their molecular mechanisms of function and biological significance remain controversial. Here, we present a genome-wide functional annotation strategy for lncRNAs based on integration of three distinct types of approaches: co-expression analysis, mapping of lncRNA-chromatin interactions and assaying molecular effects of lncRNA knockdowns obtained using an inducible and highly specific CRISPR/Cas13 system. We apply the strategy to annotate 407 vlincRNAs belonging to a novel widespread subclass of lncRNAs. We show that vlincRNAs appear to regulate, positively and negatively and both in trans and cis, multiple genes encoding proteins predominantly involved in RNA- and development-related functions, cell cycle and cellular adhesion via a potential mechanism involving proximity in nucleus. Finally, we show vlincRNAs and their regulatory networks potentially represent novel components of DNA damage response. Overall design: Regulatory networks for vlincRNAs were established based on co-expression analysis using 64 RNA-seq samples obtained using single-molecule sequencing (SMS, Helicos/SeqLL platform) from total RNA isolated from K562 cells perturbed with various drug treatments.