Dataset Information

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

ABSTRACT:

Background

The majority of the human genome is transcribed in the form of long non-coding (lnc) RNAs. While these transcripts have attracted considerable interest, their molecular mechanisms of function and biological significance remain controversial. One of the main reasons behind this lies in the significant challenges posed by lncRNAs requiring the development of novel methods and concepts to unravel their functionality. Existing methods often lack cross-validation and independent confirmation by different methodologies and therefore leave significant ambiguity as to the authenticity of the outcomes. Nonetheless, despite all the caveats, it appears that lncRNAs may function, at least in part, by regulating other genes via chromatin interactions. Therefore, the function of a lncRNA could be inferred from the function of genes it regulates. In this work, we present a genome-wide functional annotation strategy for lncRNAs based on identification of their regulatory networks via the 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.

Results

We applied the strategy to annotate 407 very long intergenic non-coding (vlinc) RNAs belonging to a novel widespread subclass of lncRNAs. We show that vlincRNAs indeed appear to regulate multiple genes encoding proteins predominantly involved in RNA- and development-related functions, cell cycle, and cellular adhesion via a mechanism involving proximity between vlincRNAs and their targets in the nucleus. A typical vlincRNAs can be both a positive and negative regulator and regulate multiple genes both in trans and cis. Finally, we show vlincRNAs and their regulatory networks potentially represent novel components of DNA damage response and are functionally important for the ability of cancer cells to survive genotoxic stress.

Conclusions

This study provides strong evidence for the regulatory role of the vlincRNA class of lncRNAs and a potentially important role played by these transcripts in the hidden layer of RNA-based regulation in complex biological systems.

SUBMITTER: Cao H

PROVIDER: S-EPMC8139166 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundLong intergenic non-coding RNAs (lincRNAs) can act as regulators of expression of protein-coding genes. Trans-natural antisense transcripts (trans-NATs) are a type of lincRNAs that contain sequence complementary to mRNA from other loci. The regulatory potential of trans-NATs has been poorly studied in eukaryotes and no example of trans-NATs regulating gene expression in plants are reported. The goal of this study was to identify lincRNAs, and particularly trans-NATs, in Arabidopsis thaliana that have a potential to regulate expression of target genes in trans at the transcriptional or translational level.ResultsWe identified 1001 lincRNAs using an RNAseq dataset from total polyA+ and polysome-associated RNA of seedlings grown under high and low phosphate, or shoots and roots treated with different phytohormones, of which 550 were differentially regulated. Approximately 30% of lincRNAs showed conservation amongst Brassicaceae and 25% harbored transposon element (TE) sequences. Gene co-expression network analysis highlighted a group of lincRNAs associated with the response of roots to low phosphate. A total of 129 trans-NATs were predicted, of which 88 were significantly differentially expressed under at least one pairwise comparison. Five trans-NATs showed a positive correlation between their expression and target mRNA steady-state levels, and three showed a negative correlation. Expression of four trans-NATs positively correlated with a change in target mRNA polysome association. The regulatory potential of these trans-NATs did not implicate miRNA mimics nor siRNAs. We also looked for lincRNAs that could regulate gene expression in trans by Watson-Crick DNA:RNA base pairing with target protein-encoding loci. We identified 100 and 81 with a positive or negative correlation, respectively, with steady-state level of their predicted target. The regulatory potential of one such candidate lincRNA harboring a SINE TE sequence was validated in a protoplast assay on three distinct genes containing homologous TE sequence in their promoters. Construction of networks highlighted other putative lincRNAs with multiple predicted target loci for which expression was positively correlated with target gene expression.ConclusionsThis study identified lincRNAs in Arabidopsis with potential in regulating target gene expression in trans by both RNA:RNA and RNA:DNA base pairing and highlights lincRNAs harboring TE sequences in such activity.

Project description:BACKGROUND:Long non-coding RNAs (lncRNAs) are key regulators of diverse cellular processes. Although a number of studies have reported the identification of bovine lncRNAs across many tissues, very little is known about the identity and characteristics of lncRNAs in bovine oocytes. METHODS:A bovine oocyte cDNA library was constructed and sequenced using the Illumina HiSeq 2000 sequencing system. The oocyte transcriptome was constructed using the ab initio assembly software Scripture and Cufflinks. The assembled transcripts were categorized to identify the novel intergenic transcripts, and the coding potential of these novel transcripts was assessed using CPAT and PhyloCSF. The resulting candidate long intergenic non-coding RNAs (lincRNAs) transcripts were further evaluated to determine if any of them contain any known protein coding domains in the Pfam database. RT-PCR was used to analyze the expression of oocyte-expressed lincRNAs in various bovine tissues. RESULTS:A total of 85 million raw reads were generated from sequencing of the bovine oocyte library. Transcriptome reconstruction resulted in the assembly of a total of 42,396 transcripts from 37,678 genomic loci. Analysis of the assembled transcripts using the step-wide pipeline resulted in the identification of 1535 oocyte lincRNAs corresponding to 1183 putative non-coding genes. A comparison of the oocyte lincRNAs with the lncRNAs reported in other bovine tissues indicated that 970 of the 1535 oocyte lincRNAs appear to be unique to bovine oocytes. RT-PCR analysis of 5 selected lincRNAs showed either specific or predominant expression of 4 lincRNAs in the fetal ovary. Functional prediction of the oocyte-expressed lincRNAs suggested their involvement in oogenesis through regulating their neighboring protein-coding genes. CONCLUSIONS:This study provides a starting point for future research aimed at understanding the roles of lncRNAs in controlling oocyte development and early embryogenesis in cattle.