SHIFTR enables the unbiased and multiplexed identification of proteins bound to specific RNA regions in live cells DOI Creative Commons
Jens Aydin, Alexander Gabel, Sebastian Zielinski

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: Oct. 10, 2023

ABSTRACT RNA-protein interactions determine the cellular fate of RNA and are central to regulating gene expression outcomes in health disease. To date, no method exists that is able identify proteins interact with specific regions within endogenous RNAs live cells. Here, we develop SHIFTR ( S elective RNase H -mediated interactome framing for target regions), an efficient scalable approach bound selected using mass spectrometry. Compared state-of-the-art techniques, superior accuracy, captures close zero background requires orders magnitude lower input material. We establish workflows targeting classes different length abundance, including short long non-coding RNAs, as well mRNAs demonstrate compatible sequentially mapping interactomes multiple a single experiment. Using SHIFTR, comprehensively cis -regulatory elements located at 5ʹ 3ʹ- terminal authentic SARS-CoV-2 genome infected cells accurately recover known novel linked function these viral elements. enables systematic region-resolved any cell type has potential revolutionize our understanding transcriptomes their regulation.

Language: Английский

SND1 binds SARS-CoV-2 negative-sense RNA and promotes viral RNA synthesis through NSP9 DOI Creative Commons
Nora Schmidt, Sabina Ganskih, Yuanjie Wei

et al.

Cell, Journal Year: 2023, Volume and Issue: 186(22), P. 4834 - 4850.e23

Published: Oct. 1, 2023

Regulation of viral RNA biogenesis is fundamental to productive SARS-CoV-2 infection. To characterize host RNA-binding proteins (RBPs) involved in this process, we biochemically identified bound genomic and subgenomic RNAs. We find that the protein SND1 binds 5' end negative-sense required for synthesis. SND1-depleted cells form smaller replication organelles display diminished virus growth kinetics. discover NSP9, a RBP direct interaction partner, covalently linked ends positive- RNAs produced during These linkages occur at replication-transcription initiation sites, consistent with NSP9 priming Mechanistically, remodels occupancy alters covalent linkage initiating nucleotides RNA. Our findings implicate synthesis unravel an unsuspected role cellular orchestrating production.

Language: Английский

Citations

29
Structural and functional insights into the enzymatic plasticity of the SARS-CoV-2 NiRAN domain DOI Creative Commons
Gabriel I. Small, Olga Fedorova, Paul Dominic B. Olinares

et al.

Molecular Cell, Journal Year: 2023, Volume and Issue: 83(21), P. 3921 - 3930.e7

Published: Oct. 26, 2023

Language: Английский

Citations

9
Targeting cap1 RNA methyltransferases as an antiviral strategy DOI Creative Commons
Yuta Tsukamoto, Manabu Igarashi, Hiroki Kato

et al.

Cell chemical biology, Journal Year: 2023, Volume and Issue: 31(1), P. 86 - 99

Published: Dec. 12, 2023

Methylation is one of the critical modifications that regulates numerous biological processes. Guanine capping and methylation at 7

Language: Английский

Citations

6
SHIFTR enables the unbiased identification of proteins bound to specific RNA regions in live cells DOI Creative Commons
Jens Aydin, Alexander Gabel, Sebastian Zielinski

et al.

Nucleic Acids Research, Journal Year: 2024, Volume and Issue: 52(5), P. e26 - e26

Published: Jan. 28, 2024

Abstract RNA-protein interactions determine the cellular fate of RNA and are central to regulating gene expression outcomes in health disease. To date, no method exists that is able identify proteins interact with specific regions within endogenous RNAs live cells. Here, we develop SHIFTR (Selective RNase H-mediated interactome framing for target regions), an efficient scalable approach bound selected using mass spectrometry. Compared state-of-the-art techniques, superior accuracy, captures minimal background requires orders magnitude lower input material. We establish workflows targeting classes different length abundance, including short long non-coding RNAs, as well mRNAs demonstrate compatible sequentially mapping interactomes multiple a single experiment. Using SHIFTR, comprehensively cis-regulatory elements located at 5′ 3′-terminal authentic SARS-CoV-2 infected cells accurately recover known novel linked function these viral elements. enables systematic region-resolved any cell type has potential revolutionize our understanding transcriptomes their regulation.

Language: Английский

Citations

2
Viral RNA capping: Mechanisms and antiviral therapy DOI

Saini Chen,

Zhimin Jiang,

Qiuchen Li

et al.

Journal of Medical Virology, Journal Year: 2024, Volume and Issue: 96(5)

Published: April 29, 2024

Abstract RNA capping is an essential trigger for protein translation in eukaryotic cells. Many viruses have evolved various strategies initiating the of viral genes and generating progeny virions infected cells via synthesizing cap structure or stealing from nascent host messenger ribonucleotide acid (mRNA). In addition to translation, a new understanding role antiviral innate immunity has advanced field mRNA synthesis vitro therapeutic applications. Recent studies on these systems revealed startlingly diverse ways molecular machinery. A comprehensive how accomplish pivotal designing effective broad‐spectrum therapies. Here we systematically review contemporary insights into RNA‐capping mechanisms employed by causing human animal infectious diseases, while also highlighting its impact immune response. The applications targeting against infections development inhibitors are summarized.

Language: Английский

Citations

1
Viral RNA Interactome: The Ultimate Researcher’s Guide to RNA–Protein Interactions DOI Creative Commons

W. B. Hanson,

Gabriel A. Romero Agosto,

Silvi Rouskin

et al.

Viruses, Journal Year: 2024, Volume and Issue: 16(11), P. 1702 - 1702

Published: Oct. 30, 2024

RNA molecules in the cell are bound by a multitude of RNA-binding proteins (RBPs) with variety regulatory consequences. Often, interactions these facilitated complex secondary and tertiary structures molecules. Viral RNAs especially known to be heavily structured interact many RBPs, roles including genome packaging, immune evasion, enhancing replication transcription, increasing translation efficiency. As such, RNA-protein interactome represents critical facet viral cycle. Characterization is necessary for development novel therapeutics targeted at disruption essential cycle events. In this review, we aim summarize various shaping interactome, interactions, as well up-to-date methods developed characterization directions novel, RNA-directed therapeutics.

Language: Английский

Citations

0
SHIFTR enables the unbiased and multiplexed identification of proteins bound to specific RNA regions in live cells DOI Creative Commons
Jens Aydin, Alexander Gabel, Sebastian Zielinski

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: Oct. 10, 2023

ABSTRACT RNA-protein interactions determine the cellular fate of RNA and are central to regulating gene expression outcomes in health disease. To date, no method exists that is able identify proteins interact with specific regions within endogenous RNAs live cells. Here, we develop SHIFTR ( S elective RNase H -mediated interactome framing for target regions), an efficient scalable approach bound selected using mass spectrometry. Compared state-of-the-art techniques, superior accuracy, captures close zero background requires orders magnitude lower input material. We establish workflows targeting classes different length abundance, including short long non-coding RNAs, as well mRNAs demonstrate compatible sequentially mapping interactomes multiple a single experiment. Using SHIFTR, comprehensively cis -regulatory elements located at 5ʹ 3ʹ- terminal authentic SARS-CoV-2 genome infected cells accurately recover known novel linked function these viral elements. enables systematic region-resolved any cell type has potential revolutionize our understanding transcriptomes their regulation.

Language: Английский

Citations

0