ARTDeco: automatic readthrough transcription detection

Samuel J Roth,Sven Heinz,Christopher Benner

doi:10.1186/s12859-020-03551-0

Samuel J Roth, Sven Heinz + Show 1 more

Open Access

https://doi.org/10.1186/s12859-020-03551-0

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Journal: BMC bioinformatics	Publication Date: May 26, 2020
Citations: 29	License type: open-access

Affiliation: University of California, San Diego

Abstract

BackgroundMounting evidence suggests several diseases and biological processes target transcription termination to misregulate gene expression. Disruption of transcription termination leads to readthrough transcription past the 3′ end of genes, which can result in novel transcripts, changes in epigenetic states and altered 3D genome structure.ResultsWe developed Automatic Readthrough Transcription Detection (ARTDeco), a tool to detect and analyze multiple features of readthrough transcription from RNA-seq and other next-generation sequencing (NGS) assays that profile transcriptional activity. ARTDeco robustly quantifies the global severity of readthrough phenotypes, and reliably identifies individual genes that fail to terminate (readthrough genes), are aberrantly transcribed due to upstream termination failure (read-in genes), and novel transcripts created as a result of readthrough (downstream of gene or DoG transcripts). We used ARTDeco to characterize readthrough transcription observed during influenza A virus (IAV) infection, validating its specificity and sensitivity by comparing its performance in samples infected with a mutant virus that fails to block transcription termination. We verify ARTDeco’s ability to detect readthrough as well as identify read-in genes from different experimental assays across multiple experimental systems with known defects in transcriptional termination, and show how these results can be leveraged to improve the interpretation of gene expression and downstream analysis. Applying ARTDeco to a gene expression data set from IAV-infected monocytes from different donors, we find strong evidence that read-in gene-associated expression quantitative trait loci (eQTLs) likely regulate genes upstream of read-in genes. This indicates that taking readthrough transcription into account is important for the interpretation of eQTLs in systems where transcription termination is blocked.ConclusionsARTDeco aids researchers investigating readthrough transcription in a variety of systems and contexts.

Highlights

Mounting evidence suggests several diseases and biological processes target transcription termination to misregulate gene expression
Automatic Readthrough Transcription Detection (ARTDeco) processes next-generation sequencing (NGS) data (e.g., RNA-seq) to characterize the features of readthrough transcription genome-wide. This includes the identification of genes that exhibit transcription downstream of their 3′ ends, genes that are transcribed as a result of readthrough transcription from upstream genes, as well as detection of novel Downstream of Gene (DoG) transcripts created as a result of readthrough transcription
The intervals used to calculate intergenic transcription levels exclude regions immediately upstream of the transcription start site (TSS, > 1 kb) and downstream of the transcription termination site (TTS, > 10 kb) to avoid detection of RNA signal that arises from incorrect Transcription start site (TSS) assignment and post-poly(A) site cleavage transcripts that may accumulate during normal termination, respectively

Summary

Introduction

Mounting evidence suggests several diseases and biological processes target transcription termination to misregulate gene expression. Transcription termination is a fundamental step in gene expression regulation. Transcription termination is triggered when RNA polymerase II (RNAPII) transcribes a polyadenylation site (PAS) that activates the cleavage and polyadenylation (CPA) complex associated with the C-terminal domain (CTD) of RNAPII [13]. There are two popular models for how CPA recruitment induces transcription termination. Recruitment of CPA is accompanied by a conformational change in elongating RNAPII, causing dissociation from the DNA and release of the nascent pre-mRNA [36]. PolyA-dependent cleavage of pre-mRNA by CPA leaves an uncapped nascent RNA emanating from elongating RNAPII. The exonuclease XRN2 degrades the unprotected nascent transcript until it catches up to transcribing RNAPII, causing its release from the DNA [12, 34]. Alternative transcription termination mechanisms have been described for histone genes, snRNAs, and transcripts generated by RNAPI and RNAPIII [11, 21, 25]

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