Evidence map›Paper›PMID 41282130›Full record

ArticleResearch square2025

A combined PIE-FRET and FCS assay to monitor RNA dynamics and cleavage by SARS-CoV-2 Nsp15.

Kenya Gordon, Zoe Wright, Meredith Frazier, Benjamin S Clark, Isha Wilson, Cameron R Stephens, Irene Silvernail, Robin E Stanley, Sharonda J LeBlanc

Abstract readPreprint
PubMed Full text at PMC Publisher
In one paragraph

Article in Research square, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

0numbers the graph read from it
0cells of the map it votes in
0citing papers in PubMed
–field-weighted citation impact
1 · What the graph read from it

What it found

Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.

The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.

2 · The registry

The trial behind it

Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.

Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.

3 · Its place in the literature

Who cites it

0 citing papers in PubMed.

No citing paper in PubMed yet.

4 · The record

Corrections and comments

5 · Who and what money

Authors and funding

9 authors.

Kenya GordonDepartment of Physics, North Carolina State University, Raleigh, NC 27695-8202, USA.
Zoe WrightMolecular and Cellular Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, 111 T. W. Alexander Drive, Research Triangle Park, NC 27709, USA.
Meredith FrazierMolecular and Cellular Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, 111 T. W. Alexander Drive, Research Triangle Park, NC 27709, USA.
Benjamin S ClarkDepartment of Physics, North Carolina State University, Raleigh, NC 27695-8202, USA.
Isha WilsonMolecular and Cellular Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, 111 T. W. Alexander Drive, Research Triangle Park, NC 27709, USA.
Cameron R StephensDepartment of Physics, North Carolina State University, Raleigh, NC 27695-8202, USA.
Irene SilvernailDepartment of Physics, North Carolina State University, Raleigh, NC 27695-8202, USA.
Robin E StanleyMolecular and Cellular Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, 111 T. W. Alexander Drive, Research Triangle Park, NC 27709, USA.
Sharonda J LeBlancDepartment of Physics, North Carolina State University, Raleigh, NC 27695-8202, USA.

Funding

Structural and Functional Characterization of the SARS-CoV-2 Endoribonuclease Nsp15ZIAES103340 · NIEHS · NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES · PI STANLEY, ROBIN · 2020 to 2025
$3.6M
Intramural NIH HHS ZIA ES103340
6 · The paper itself

Abstract

Nsp15 is an endoribonuclease, highly conserved among coronaviruses, that helps the virus evade detection in host cells by cleaving uridine-rich viral RNA sequences. Those sequences would otherwise trigger immune response pathways. Its essential role in pathogenesis and highly conserved nature make Nsp15 an attractive target for therapeutic intervention. While its crystal structure and uridine specificity are well-established, the influence of RNA structural dynamics and divalent cations on Nsp15 activity remains less understood. Leveraging single-molecule Pulsed Interleaved Excitation (PIE)-FRET and Fluorescence Correlation Spectroscopy (FCS) in combination, we developed an assay to track RNA cleavage by Nsp15 variants in real time and monitor the conformational dynamics of hybrid RNA/DNA substrates. Using our methodology and analysis strategies, we obtained clear indicators RNA cleavage with both PIE-FRET and FCS data analysis. Our assay also revealed signatures of unique dynamic behavior in uridine-containing RNA substrates, indicating that divalent cations enhance substrate flexibility, which increases the reaction rate of RNA cleavage by Nsp15. We propose that divalent cations induce a more favorable conformation in single-stranded RNA substrates that mediate the efficient nuclease activity of Nsp15.

Indexed as

FCSFRETNsp15RNA dynamicssingle-molecule

Identifiers

PMID41282130
PMCPMC12632706

What OpenQuestion holds

Textmetadata
LicenceCC BY
Read underepoch 390

Registered trials

None linked

Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.