A pan-cancer transcriptome analysis of exitron splicing identifies novel cancer driver genes and neoepitopes

Wang, Ting You; Liu, Qi; Ren, Yanan; Alam, Sk Kayum; Wang, Li; Zhu, Zhu; Hoeppner, Luke H.; Dehm, Scott M.; Cao, Qi; Yang, Rendong

doi:10.1016/j.molcel.2021.03.028

Cited by 49 publications

(48 citation statements)

References 88 publications

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“…2 f). This is potentially important, because a recent paper reported more than 100,000 exitrons in the TCGA database and suggested that the corresponding isoforms are novel cancer drivers and neoepitopes [ 23 ]. To learn whether such analyses might be affected by RT artifacts, we overlaid the falsitrons from our ONT data comparison onto these reported exitrons.…”

Section: Resultsmentioning

confidence: 99%

“…The k -mers were required to overlap at least 1 nt of the 5′ and 3′ dinucleotide motifs. The same analysis was applied to all the exitrons detected in Wang et al [ 23 ] as well as for all unique annotated introns in GENCODE gene annotation (v36, genome version hg38) [ 33 ].…”

Section: Methodsmentioning

confidence: 99%

“…For the sequence logos at splice sites, we retrieved the sequence in a 15-nt window (3 nt in the exon + 12 nt in the intron) of the 3′ and 5′ splice sites of the different sets of introns: our putative falsitrons from the ONT comparison ( n = 57), all unique exitrons reported by Wang et al [ 23 ] ( n = 123,337) and all unique introns in GENCODE gene annotation (v36, genome version hg38) ( n = 387,483). We used the R package ggseqlogo [ 37 ] to plot the frequency of nucleotides in each set.…”

Section: Methodsmentioning

confidence: 99%

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Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts

et al. 2021

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Resistance to CD19-directed immunotherapies in lymphoblastic leukemia has been attributed, among other factors, to several aberrant CD19 pre-mRNA splicing events, including recently reported excision of a cryptic intron embedded within CD19 exon 2. While “exitrons” are known to exist in hundreds of human transcripts, we discovered, using reporter assays and direct long-read RNA sequencing (dRNA-seq), that the CD19 exitron is an artifact of reverse transcription. Extending our analysis to publicly available datasets, we identified dozens of questionable exitrons, dubbed “falsitrons,” that appear only in cDNA-seq, but never in dRNA-seq. Our results highlight the importance of dRNA-seq for transcript isoform validation.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts

et al. 2021

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“…Notably, a series of intronic DNA variations have been reported to affect gene expression, RNA splicing, and to cause cancers and nervous system disorders [32][33][34][35][36] . Such risks accompany GEIS and may affect experiments.…”

Section: Discussionmentioning

confidence: 99%

Efficient Gene Editing Through an Intronic Selection Marker in Cells

Wang

et al. 2021

Preprint

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Background Gene editing technology has provided researchers with the ability to modify genome sequences in almost all eukaryotes. Gene-edited cell lines are being used with increasing frequency in both bench research and targeted therapy. Despite the great importance and universality of gene editing, however, precision and efficiency are hard to achieve with the prevailing editing strategies, such as homology-directed DNA repair (HDR) and the use of base editors (BEs). Results & Discussion Our group has developed a novel gene editing technology to indicate DNA variation with an independent selection marker using an HDR strategy, which we named Gene Editing through an Intronic Selection marker (GEIS). GEIS uses a simple process to avoid nonhomologous end joining (NHEJ)-mediated false-positive effects and achieves editing efficiency as high as 91% without disturbing endogenous gene splicing and expression. We re-examined the correlation of the conversion tract and editing efficiency, and our data suggest that GEIS has the potential to edit approximately 99% of gene editing targets in human and mouse cells. The results of further comprehensive analysis suggest that the strategy may be useful for introducing multiple DNA variations in cells.

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“…The protocol constitutes a step-by-step guide from data collection to neoantigen prediction. For complete details on the use and execution of this protocol, please refer to Wang et al. (2021) .…”

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confidence: 99%

Integrated protocol for exitron and exitron-derived neoantigen identification using human RNA-seq data with ScanExitron and ScanNeo

Wang

Yang

2021

STAR Protocols

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Summary Exitron splicing (EIS) events in cancers can disrupt functional protein domains to cause cancer driver effects. EIS has been recognized as a new source of tumor neoantigens. Here, we describe an integrated protocol for EIS and EIS-derived neoantigen identification using RNA-seq data. The protocol constitutes a step-by-step guide from data collection to neoantigen prediction. For complete details on the use and execution of this protocol, please refer to Wang et al. (2021) .

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A pan-cancer transcriptome analysis of exitron splicing identifies novel cancer driver genes and neoepitopes

Cited by 49 publications

References 88 publications

Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts

Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts

Efficient Gene Editing Through an Intronic Selection Marker in Cells

Integrated protocol for exitron and exitron-derived neoantigen identification using human RNA-seq data with ScanExitron and ScanNeo

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