Long-Read cDNA Sequencing Enables a “Gene-Like” Transcript Annotation of Transposable Elements

Panda, Kaushik; Slotkin, R. Keith

doi:10.1105/tpc.20.00115

Cited by 51 publications

(65 citation statements)

References 48 publications

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“…Although some tools have been developed so far to overcome these obstacles, most of the lncRNA identification pipelines still ignore ambiguously mapped reads [ 24 , 25 ]. Thus, long-read sequencing technologies provide a great opportunity for transcriptome exploration, including the identification of transcribed repeats (e.g., transposable elements) or repeat-related transcripts [ 26 ]. Moreover, Panda and Slotkin [ 26 ] showed that by using Nanopore long cDNA reads, it is even possible to trace the expression of individual TEs from multicopy families in Arabidopsis and maize.…”

Section: Introductionmentioning

confidence: 99%

“…Thus, long-read sequencing technologies provide a great opportunity for transcriptome exploration, including the identification of transcribed repeats (e.g., transposable elements) or repeat-related transcripts [ 26 ]. Moreover, Panda and Slotkin [ 26 ] showed that by using Nanopore long cDNA reads, it is even possible to trace the expression of individual TEs from multicopy families in Arabidopsis and maize. The application of long-read technologies to the exploration of the lncRNA repertoire in plants has been demonstrated for several plant species, including Oryza sativa L. ssp.…”

Section: Introductionmentioning

confidence: 99%

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Nanopore RNA Sequencing Revealed Long Non-Coding and LTR Retrotransposon-Related RNAs Expressed at Early Stages of Triticale SEED Development

Kirov

Dudnikov

Merkulov³

et al. 2020

Plants

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The intergenic space of plant genomes encodes many functionally important yet unexplored RNAs. The genomic loci encoding these RNAs are often considered “junk”, DNA as they are frequently associated with repeat-rich regions of the genome. The latter makes the annotations of these loci and the assembly of the corresponding transcripts using short RNAseq reads particularly challenging. Here, using long-read Nanopore direct RNA sequencing, we aimed to identify these “junk” RNA molecules, including long non-coding RNAs (lncRNAs) and transposon-derived transcripts expressed during early stages (10 days post anthesis) of seed development of triticale (AABBRR, 2n = 6x = 42), an interspecific hybrid between wheat and rye. Altogether, we found 796 lncRNAs and 20 LTR retrotransposon-related transcripts (RTE-RNAs) expressed at this stage, with most of them being previously unannotated and located in the intergenic as well as intronic regions. Sequence analysis of the lncRNAs provide evidence for the frequent exonization of Class I (retrotransposons) and class II (DNA transposons) transposon sequences and suggest direct influence of “junk” DNA on the structure and origin of lncRNAs. We show that the expression patterns of lncRNAs and RTE-related transcripts have high stage specificity. In turn, almost half of the lncRNAs located in Genomes A and B have the highest expression levels at 10–30 days post anthesis in wheat. Detailed analysis of the protein-coding potential of the RTE-RNAs showed that 75% of them carry open reading frames (ORFs) for a diverse set of GAG proteins, the main component of virus-like particles of LTR retrotransposons. We further experimentally demonstrated that some RTE-RNAs originate from autonomous LTR retrotransposons with ongoing transposition activity during early stages of triticale seed development. Overall, our results provide a framework for further exploration of the newly discovered lncRNAs and RTE-RNAs in functional and genome-wide association studies in triticale and wheat. Our study also demonstrates that Nanopore direct RNA sequencing is an indispensable tool for the elucidation of lncRNA and retrotransposon transcripts.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Nanopore RNA Sequencing Revealed Long Non-Coding and LTR Retrotransposon-Related RNAs Expressed at Early Stages of Triticale SEED Development

Kirov

Dudnikov

Merkulov³

et al. 2020

Plants

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show abstract

“…The vast majority of previous studies of RTE expression used short RNAseq reads to identify expressed RTEs. Although RNAseq is a robust method of gene expression analysis, it is challenging to estimate retrotransposon expression using this approach [8,25,48]. Our first attempt to detect expressed RTEs by RNAseq led to the identification of >300 expressed RTEs.…”

Section: Rnaseq and Nanopore Rna Sequencing Are Complementary Approacmentioning

confidence: 99%

“…No algorithms exist that reconstruct entire TE transcripts. Nanopore RNA (cDNA) sequencing has recently been applied to plant TE "gene-like" annotation in an Arabidopsis mutant defective in the main TE-silencing pathways [25]. The nanopore cDNA reads from Arabidopsis and the PacBio reads from maize can be exploited to precisely determine TE transcripts and transcription features, including transcription start sites and poly-A tail length.…”

Section: Introductionmentioning

confidence: 99%

Genomic and Transcriptomic Survey Provides New Insight into the Organization and Transposition Activity of Highly Expressed LTR Retrotransposons of Sunflower (Helianthus annuus L.)

Kirov

Omarov

Merkulov

et al. 2020

IJMS

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LTR retrotransposons (RTEs) play a crucial role in plant genome evolution and adaptation. Although RTEs are generally silenced in somatic plant tissues under non-stressed conditions, some expressed RTEs (exRTEs) escape genome defense mechanisms. As our understanding of exRTE organization in plants is rudimentary, we systematically surveyed the genomic and transcriptomic organization and mobilome (transposition) activity of sunflower (Helianthus annuus L.) exRTEs. We identified 44 transcribed RTEs in the sunflower genome and demonstrated their distinct genomic features: more recent insertion time, longer open reading frame (ORF) length, and smaller distance to neighboring genes. We showed that GAG-encoding ORFs are present at significantly higher frequencies in exRTEs, compared with non-expressed RTEs. Most exRTEs exhibit variation in copy number among sunflower cultivars and one exRTE Gagarin produces extrachromosomal circular DNA in seedling, demonstrating recent and ongoing transposition activity. Nanopore direct RNA sequencing of full-length RTE RNA revealed complex patterns of alternative splicing in RTE RNAs, resulting in isoforms that carry ORFs for distinct RTE proteins. Together, our study demonstrates that tens of expressed sunflower RTEs with specific genomic organization shape the hidden layer of the transcriptome, pointing to the evolution of specific strategies that circumvent existing genome defense mechanisms.

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“…In a new study, Panda and Slotkin used Oxford Nanopore Technology long-read sequencing of cDNAs to establish a transcriptbased annotation of TEs in Arabidopsis thaliana (Panda and Slotkin, 2020) ( Figure 1A). To expose TE transcripts for sequencing and annotation, the authors used a combination of Arabidopsis mutants compromised in multiple layers of TE silencing.…”

Section: In the Transcripts: Long-read Transcriptomics Enables A Novementioning

confidence: 99%

In the Transcripts: Long-Read Transcriptomics Enables a Novel Type of Transposable Element Annotation in Plants

Benoit

2020

The Plant Cell

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Long-Read cDNA Sequencing Enables a “Gene-Like” Transcript Annotation of Transposable Elements

Abstract: A transcript-based annotation of transposable elements allows for reduced bioinformatic complexity and direct hypothesis testing of previously challenging regions of the genome.

Cited by 51 publications

References 48 publications

Nanopore RNA Sequencing Revealed Long Non-Coding and LTR Retrotransposon-Related RNAs Expressed at Early Stages of Triticale SEED Development

Nanopore RNA Sequencing Revealed Long Non-Coding and LTR Retrotransposon-Related RNAs Expressed at Early Stages of Triticale SEED Development

Genomic and Transcriptomic Survey Provides New Insight into the Organization and Transposition Activity of Highly Expressed LTR Retrotransposons of Sunflower (Helianthus annuus L.)

In the Transcripts: Long-Read Transcriptomics Enables a Novel Type of Transposable Element Annotation in Plants

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