Sequence determinants of polyadenylation-mediated regulation

Slutskin, Ilya Vainberg; Weinberger, Adina; Segal, Eran

doi:10.1101/gr.247312.118

Cited by 22 publications

(23 citation statements)

References 71 publications

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“…In a recent study, a polyadenylation efficiency prediction tool (polyApredictor) was developed based on a deep learning model (Vainberg Slutskin et al 2019). We therefore conducted a comparison of the performance between our mpCHECK2 system and polyApredictor.…”

Section: High Correlation Of the Performance Between Mpcheck2 And Polyapredictor In Evaluating The Effect Of Pas Variants On Gene Expressmentioning

confidence: 99%

Systematic evaluation of the effect of polyadenylation signal variants on the expression of disease-associated genes

Chen

Wei

et al. 2021

Genome Res.

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Single nucleotide variants (SNVs) within polyadenylation signals (PASs), a specific six-nucleotide sequence required for mRNA maturation, can impair RNA-level gene expression and cause human diseases. However, there is a lack of genome-wide investigation and systematic confirmation tools for identifying PAS variants. Here, we present a computational strategy to integrate the most reliable resources for discovering distinct genomic features of PAS variants and also develop a credible and convenient experimental tool to validate the effect of PAS variants on expression of disease-associated genes. This approach will greatly accelerate the deciphering of PAS variation-related human diseases.

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Section: High Correlation Of the Performance Between Mpcheck2 And Polyapredictor In Evaluating The Effect Of Pas Variants On Gene Expressmentioning

confidence: 99%

Systematic evaluation of the effect of polyadenylation signal variants on the expression of disease-associated genes

Chen

Wei

et al. 2021

Genome Res.

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“…The cloning steps were performed essentially as described previously (Vainberg Slutskin et al, 2018). 714…”

Section: Synthetic Library Cloning 713mentioning

confidence: 99%

High-throughput interrogation of programmed ribosomal frameshifting in human cells

Mikl

Pilpel

Segal

2018

Preprint

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Programmed ribosomal frameshifting is the controlled slippage of the translating ribosome to an alternative frame. This tightly regulated process is widely employed by human viruses such as HIV and SARS-CoV and is critical for their life cycle and virulence. It is also utilized throughout the tree of life to implement a feedback control mechanism to regulate polyamine levels. However, despite its universality and clinical relevance, a limited number of studies investigated this process on only a few selected examples, largely due to a lack of experimental means. Here, we developed a highthroughput, fluorescence-based approach to assay the frameshifting potential of a sequence. We designed and tested >12.000 sequences based on 15 viral and human frameshifting events, allowing us to elucidate the rules governing ribosomal frameshifting in a systematic way and to discover novel regulatory features. We also utilized our approach to search for novel frameshifting events and identified dozens of previously unknown frameshifting sites in human, showing that programmed ribosomal frameshifting is more common than previously anticipated. We assessed the natural variation in HIV gag-pol frameshifting rates by testing >500 clinical isolates and identified subtypespecific differences as well as associations between viral load in patients and the optimality of gagpol frameshifting rates. We further devised a machine learning algorithm that accurately predicts frameshifting rates of novel variants (up to r=0.70), including subtle differences between HIV isolates (r=0.44), providing a basis for the development of antiviral agents acting on programmed ribosomal frameshifting. author/funder. All rights reserved. No reuse allowed without permission.

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“…Large scale testing of rationally designed or random sequence libraries has immensely contributed to elucidating the regulatory grammar of transcription (Arnold et al, 2013;Grossman et al, 2017;Kheradpour et al, 2013;Patwardhan et al, 2012;Sharon et al, 2012) , splicing (Adamson et al, 2018;Mikl et al, 2019;Rosenberg et al, 2015;Soemedi et al, 2017;Wong et al, 2018) , polyadenylation (Bogard et al, 2019;Vainberg Slutskin et al, 2019), miRNA-mediated regulation (Slutskin et al, 2018), other forms of translational control (Mikl et al, 2020;Weingarten-Gabbay et al, 2016) , and RNA nuclear enrichment and export (Lubelsky and Ulitsky, 2018;Shukla et al, 2018;Zuckerman et al, 2020) demonstrating the power and universal applicability of such approaches. Although they have become an experimental pillar of studies in gene regulation, a dedicated high-throughput systematic attempt to dissect the regulatory logic of subcellular RNA localization is still lacking.…”

Section: Introductionmentioning

confidence: 99%

A massively parallel reporter assay reveals focused and broadly encoded RNA localization signals in neurons

Mikl

Eletto

Lee

et al. 2021

Preprint

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Asymmetric subcellular localization of mRNA is a common cellular phenomenon that is thought to contribute to spatial gene regulation. In highly polar neurons, subcellular transcript localization and translation are thought to enhance cellular efficiency and timely responses to external cues. Although mRNA localization has been observed in many tissues and numerous examples of the functional importance of this process exist, we still lack a systematic understanding of how the transcript sorting machinery works in a sequence-specific manner. Here, we addressed these gaps by combining subcellular transcriptomics and rationally designed sequence libraries. We developed a massively parallel reporter assay (MPRA) for mRNA localization and tested ~50,000 sequences for their ability to drive RNA localization to neurites of neuronal cell lines. By scanning the 3'UTR of >300 genes we identified many previously unknown localization regions and mapped the localization potential of endogenous sequences. Our data suggest two ways the localization potential can be encoded in the 3'UTR: focused localization motifs and broadly encoded localization potential based on small contributions. We identified sequence motifs enriched in dendritically localized transcripts and tested the potential of these motifs to affect the localization behavior of an mRNA. This assay revealed sequence elements with the ability to bias localization towards neurite as well as soma. Depletion of RNA binding proteins predicted or experimentally shown to bind these motifs abolished the effect on localization, suggesting that these motifs act by recruiting specific RNA-binding proteins. Based on our dataset we developed machine learning models that accurately predict the localization behavior of novel sequences. Testing this predictor on native mRNA sequencing data showed good agreement between predicted and observed localization potential, suggesting that the rules uncovered by our MPRA also apply to the localization of native transcripts. Applying similar systematic high-throughput approaches to other cell types will open the door for a comparative perspective on RNA localization across tissues and reveal the commonalities and differences of this crucial regulatory mechanism.

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Sequence determinants of polyadenylation-mediated regulation

Cited by 22 publications

References 71 publications

Systematic evaluation of the effect of polyadenylation signal variants on the expression of disease-associated genes

Systematic evaluation of the effect of polyadenylation signal variants on the expression of disease-associated genes

High-throughput interrogation of programmed ribosomal frameshifting in human cells

A massively parallel reporter assay reveals focused and broadly encoded RNA localization signals in neurons

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