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The RNA Atlas expands the catalog of human non-coding RNAs

ABSTRACT: The human transcriptome consists of various RNA biotypes including multiple types of non-coding RNAs (ncRNAs). Current ncRNA compendia remain incomplete partially because they are almost exclusively derived from the interrogation of small- and polyadenylated RNAs. Here, we present a more comprehensive atlas of the human transcriptome that is derived from matching polyA-, total-, and small-RNA profiles of a heterogenous collection of nearly 300 human tissues and cell lines. We report thousands of novel RNA species across all major RNA biotypes, including a hitherto poorly-cataloged class of non-polyadenylated single-exon long non-coding RNAs. In addition, we exploit intron abundance estimates from total RNA-sequencing to predict the regulatory potential of various non-coding RNAs. Our study represents a substantial expansion of the current catalogue of human ncRNAs and their regulatory interactions. All data and results are accessible through the R2 webtool and serve as a basis to further explore RNA biology and function.

ORGANISM(S): Homo sapiens

PROVIDER: GSE138734 | GEO | 2021/04/15

REPOSITORIES: GEO

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Project description:The general role of RNA in the cell beyond protein biosynthesis is only beginning to emerge with little knowledge about the structural or organizational functions of RNA. In turn, functional non-coding RNAs (ncRNAs) are often identified by their regulation or impact on cellular phenotypes. However, the huge number and diversity of ncRNAs highly complicate the task of their molecular characterization and rationalization into cellular processes. For the vast majority of ncRNAs as well as for non-coding functions of known mRNAs, the mechanisms underlying their modes of action as well as their impact on protein complexes are unknown. Thus, the greatest challenge in the field of RNA biology today is the elucidation of molecular mechanisms and functional interaction partners of RNA molecules. Since RNAs do not act alone, but often interact with specific protein partners, proteomic approaches have been developed to study RNA-binding proteins (RBPs) and aim at revealing the molecular mechanisms underlying RNA function. Large-scale studies aiming at globally identifying RNA-binding proteins principally focused on poly(A)-RNA transcripts. A notable limitation of such approaches is that many ncRNAs are not polyadenylated, so that their interaction partners were not detected in these screens. Most importantly however, the overlap between the numerous studies aiming to identify RBPs is limited and their specificity remains unclear. Moreover, the previous studies were based on the identification of RBPs omitting their participation in protein-protein complexes and more interestingly in RNA-dependent protein-protein complexes. Here, we introduce the concept of "RNA dependence" to overcome these challenges. We define a protein as RNA-dependent if its interactome (hence likely its function) depends on RNA without necessarily directly binding to RNA. Based on this new concept, we developed a proteome-wide screening approach to gain mechanistic insight into the function of RNAs - both coding and non-coding - in RNA-protein complexes and their impact on the function of such complexes.

Project description:The number of annotated protein coding genes in the genome of Caenorhabditis elegans is similar to that of other animals, but the extent of its non-protein coding transcriptome remains unknown. Expression profiling on whole genome tiling microarrays applied to a mixed stage C. elegans population verified the expression of 71% of all annotated exons. Only a small fraction (11 %) of the polyadenylated transcription is non-annotated, and appears to consist of approximately 3200 missed or alternative exons and 7800 small transcripts of unknown function (TUFs). Almost half (44%) of the detected transcriptional output is non-polyadenylated and probably not protein coding, and of this 70% overlap the boundaries of protein coding genes in a complex manner. Specific analysis of small non-polyadenylated transcripts verified 98% of all annotated small ncRNAs, and suggested that the transcriptome contains about 1,200 small (<500 nt) unannoted non-coding loci. After combining overlapping transcripts, we estimate that at least 70% of the total C. elegans genome is transcribed. Keywords: RNA fractions RNA was extracted from mixed stage wild-type N2 strain worms cultivated at 20oC according to the Trizol (Invitrogen) protocol. Small RNAs (< 500nt, SNPA sample) were isolated using a Qiagen tip (Qiagen), and the Poly(A) Purist MAG (Ambion) and MicrobExpress kits (Ambion) were adapted to remove remaining mRNAs and rRNAs (Deng et al. 2006). The enriched ncRNA pool was cloned using an adaptor-mediated library construction protocol. RNAs were dephosphorylated with calf intestine alkaline phosphatase (Fermentas), then ligated to the 3M-bM-^@M-^Y-adaptor (3AD) oligonucleotide by T4 RNA ligase (Fermentas) (He et al. 2006). Polyadenylated RNA (PA sample) was isolated from total RNA using the Poly(A) Purist MAG kit (above). Non-polyadenylated RNA (NPA sample) was prepared by removing polyadenylated RNA using the Poly(A) Purist MAG kit and rRNA using the MicrobExpress kit.

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The RNA Atlas expands the catalog of human non-coding RNAs

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