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A positive readout single transcript reporter for site-specific mRNA cleavage

Cleavage of mRNA molecules causes their rapid degradation, thereby playing an important role in regulation of gene expression and host genome defense from viruses and transposons in bacterial and eukaryotic cells. Current negative-readout, and repressor-based positive-readout reporters of mRNA degra...

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Detalles Bibliográficos
Autores principales: Kandul, Nikolay, Guo, Ming, Hay, Bruce A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522606/
https://www.ncbi.nlm.nih.gov/pubmed/28740759
http://dx.doi.org/10.7717/peerj.3602
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author Kandul, Nikolay
Guo, Ming
Hay, Bruce A.
author_facet Kandul, Nikolay
Guo, Ming
Hay, Bruce A.
author_sort Kandul, Nikolay
collection PubMed
description Cleavage of mRNA molecules causes their rapid degradation, thereby playing an important role in regulation of gene expression and host genome defense from viruses and transposons in bacterial and eukaryotic cells. Current negative-readout, and repressor-based positive-readout reporters of mRNA degradation have limitations. Here we report the development of a single transcript that acts as a positive reporter of mRNA cleavage. We show that placement of bacterial CopT and CopA hairpins into the 5′ UTR and 3′ UTR of an mRNA results in inhibition of translation of the intervening coding sequence in Drosophila. An internal poly(A) tract inserted downstream of the coding sequence stabilizes transcripts cut within the 3′ UTR. When these components are combined in a transcript in which targets sites for RNA cleavage are placed between the poly(A) tract and CopA, cleavage results in translational activation, providing a single transcript-based method of sensing mRNA cleavage with a positive readout.
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spelling pubmed-55226062017-07-24 A positive readout single transcript reporter for site-specific mRNA cleavage Kandul, Nikolay Guo, Ming Hay, Bruce A. PeerJ Bioengineering Cleavage of mRNA molecules causes their rapid degradation, thereby playing an important role in regulation of gene expression and host genome defense from viruses and transposons in bacterial and eukaryotic cells. Current negative-readout, and repressor-based positive-readout reporters of mRNA degradation have limitations. Here we report the development of a single transcript that acts as a positive reporter of mRNA cleavage. We show that placement of bacterial CopT and CopA hairpins into the 5′ UTR and 3′ UTR of an mRNA results in inhibition of translation of the intervening coding sequence in Drosophila. An internal poly(A) tract inserted downstream of the coding sequence stabilizes transcripts cut within the 3′ UTR. When these components are combined in a transcript in which targets sites for RNA cleavage are placed between the poly(A) tract and CopA, cleavage results in translational activation, providing a single transcript-based method of sensing mRNA cleavage with a positive readout. PeerJ Inc. 2017-07-20 /pmc/articles/PMC5522606/ /pubmed/28740759 http://dx.doi.org/10.7717/peerj.3602 Text en ©2017 Kandul et al. http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Bioengineering
Kandul, Nikolay
Guo, Ming
Hay, Bruce A.
A positive readout single transcript reporter for site-specific mRNA cleavage
title A positive readout single transcript reporter for site-specific mRNA cleavage
title_full A positive readout single transcript reporter for site-specific mRNA cleavage
title_fullStr A positive readout single transcript reporter for site-specific mRNA cleavage
title_full_unstemmed A positive readout single transcript reporter for site-specific mRNA cleavage
title_short A positive readout single transcript reporter for site-specific mRNA cleavage
title_sort positive readout single transcript reporter for site-specific mrna cleavage
topic Bioengineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522606/
https://www.ncbi.nlm.nih.gov/pubmed/28740759
http://dx.doi.org/10.7717/peerj.3602
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