Abstract
The prevalence of long non-coding RNAs (lncRNA) and natural antisense transcripts (NATs) has been reported in a variety of organisms. While a consensus has yet to be reached on their global importance, an increasing number of examples have been shown to be functional, regulating gene expression at the transcriptional and post-transcriptional level. Here, we use RNA sequencing data from the ABI SOLiD platform to identify lncRNA and NATs obtained from samples of the filamentous fungus Neurospora crassa grown under different light and temperature conditions. We identify 939 novel lncRNAs, of which 477 are antisense to annotated genes. Across the whole dataset, the extent of overlap between sense and antisense transcripts is large: 371 sense/antisense transcripts are complementary over 500 nts or more and 236 overlap by more than 1000 nts. Most prevalent are 3′ end overlaps between convergently transcribed sense/antisense pairs, but examples of divergently transcribed pairs and nested transcripts are also present. We confirm the expression of a subset of sense/antisense transcript pairs by qPCR. We examine the size, types of overlap and expression levels under the different environmental stimuli of light and temperature, and identify 11 lncRNAs that are up-regulated in response to light. We also find differences in transcript length and the position of introns between protein-coding transcripts that have antisense expression and transcripts with no antisense expression. These results demonstrate the ability of N. crassa lncRNAs and NATs to be regulated by different environmental stimuli and provide the scope for further investigation into the function of NATs.
Highlights
High-throughput sequencing has revealed that the overwhelming majority of the eukaryotic genome is transcribed
To eliminate the possibility that the antisense long non-coding RNAs (lncRNA) could be annotated based on reads that are mapped to the incorrect genomic strand, we examined the mapped reads for splice junctions. 315 of our antisense lncRNAs (66%) have consensus splice junctions (GT-AG, GC-AG and AT-AC) supported by reads spanning the intron
We identify 38 pairs of sense/antisense lncRNAs originating from previously unannotated loci (Table S6). 36 of these pairs overlap by more than 200 nts, and we again observe an excess of convergently transcribed sense/antisense pairs (14 of these pairs overlap at the 39 end, and 9 at the 59 end)
Summary
High-throughput sequencing has revealed that the overwhelming majority of the eukaryotic genome is transcribed. The ENCODE project has annotated transcription originating from around three quarters of the human genome [1,2,3]. The majority of the mouse genome has been shown to be transcribed [4]. Novel transcribed regions may represent extensions of known protein-coding genes, novel protein-coding transcripts, and transcripts that do not appear to have proteincoding capacity [5]. High-throughput technologies have highlighted thousands of lncRNAs in a range of eukaryotic organisms, from yeast to humans [8,9,10,11]. A handful of examples have been well-characterised and shown to have roles in the transcriptional and post-transcriptional control of gene expression via RNA-protein, RNA-DNA and RNA-RNA interactions. The function of the vast majority of lncRNAs remains a mystery
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