Evolinc: a comparative transcriptomics and genomics pipeline for quickly identifyingsequence conserved lincRNAs for functional analysis.

Abstract

Long intergenic non-coding RNAs (lincRNAs) are an abundant and functionally diverse class of eukaryotic transcripts. Reported lincRNA repertoires in mammals vary, but are commonly in the thousands to tens of thousands of transcripts, covering ~90% of the genome. In addition to elucidating function, there is particular interest in understanding the origin and evolution of lincRNAs. Aside from mammals, lincRNA populations have been sparsely sampled, precluding evolutionary analyses focused on lincRNA emergence and persistence. Here we present Evolinc, a two-module pipeline designed to facilitate lincRNA discovery and characterize aspects of lincRNA evolution. The first module (Evolinc-I) is a lincRNA identification workflow that also facilitates downstream differential expression analysis and genome browser visualization of identified lincRNAs. The second module (Evolinc-II) is a genomic and transcriptomic comparative analyses workflow that determines the phylogenetic depth to which a lincRNA locus is conserved within a user-defined group of related species. Evolinc-II builds families of homologous lincRNA loci, aligns constituent sequences, infers gene trees, and then uses gene tree / species tree reconciliation to reconstruct evolutionary processes such as gain, loss, or duplication of the locus. Here we demonstrate that Evolinc-I is agnostic to target organism by validating against previously annotated Arabidopsis and human lincRNA data. Using Evolinc-II, we examine ways in which conservation can rapidly be used to winnow down large lincRNA datasets to a small set of candidates for functional analysis. Finally, we show how Evolinc-II can be used to recover the evolutionary history of a known lincRNA, the human telomerase RNA (TERC). The analyses revealed unexpected duplication events as well as the loss and subsequent acquisition of a novel TERC locus in the lineage leading to mice and rats. The Evolinc pipeline is currently integrated in CyVerses Discovery Environment and is free to use by researchers.