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Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
The contribution of specific factors to bacterial virulence is generally investigated through creation of genetic “knockouts” that are then compared to wild-type strains or complemented mutants. This paradigm is useful to understand the effect of presence vs. absence of a specific gene product but c...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370707/ https://www.ncbi.nlm.nih.gov/pubmed/25798590 http://dx.doi.org/10.1371/journal.pone.0119823 |
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author | Amaral, Fábio E. Parker, Dane Randis, Tara M. Kulkarni, Ritwij Prince, Alice S. Shirasu-Hiza, Mimi M. Ratner, Adam J. |
author_facet | Amaral, Fábio E. Parker, Dane Randis, Tara M. Kulkarni, Ritwij Prince, Alice S. Shirasu-Hiza, Mimi M. Ratner, Adam J. |
author_sort | Amaral, Fábio E. |
collection | PubMed |
description | The contribution of specific factors to bacterial virulence is generally investigated through creation of genetic “knockouts” that are then compared to wild-type strains or complemented mutants. This paradigm is useful to understand the effect of presence vs. absence of a specific gene product but cannot account for concentration-dependent effects, such as may occur with some bacterial toxins. In order to assess threshold and dose-response effects of virulence factors, robust systems for tunable expression are required. Recent evidence suggests that the folding free energy (ΔG) of the 5’ end of mRNA transcripts can have a significant effect on translation efficiency and overall protein abundance. Here we demonstrate that rational alteration of 5’ mRNA folding free energy by introduction of synonymous mutations allows for predictable changes in pneumolysin (PLY) expression by Streptococcus pneumoniae without the need for chemical inducers or heterologous promoters. We created a panel of isogenic S. pneumoniae strains, differing only in synonymous (silent) mutations at the 5’ end of the PLY mRNA that are predicted to alter ΔG. Such manipulation allows rheostat-like control of PLY production and alters the cytotoxicity of whole S. pneumoniae on primary and immortalized human cells. These studies provide proof-of-principle for further investigation of mRNA ΔG manipulation as a tool in studies of bacterial pathogenesis. |
format | Online Article Text |
id | pubmed-4370707 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-43707072015-04-04 Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae Amaral, Fábio E. Parker, Dane Randis, Tara M. Kulkarni, Ritwij Prince, Alice S. Shirasu-Hiza, Mimi M. Ratner, Adam J. PLoS One Research Article The contribution of specific factors to bacterial virulence is generally investigated through creation of genetic “knockouts” that are then compared to wild-type strains or complemented mutants. This paradigm is useful to understand the effect of presence vs. absence of a specific gene product but cannot account for concentration-dependent effects, such as may occur with some bacterial toxins. In order to assess threshold and dose-response effects of virulence factors, robust systems for tunable expression are required. Recent evidence suggests that the folding free energy (ΔG) of the 5’ end of mRNA transcripts can have a significant effect on translation efficiency and overall protein abundance. Here we demonstrate that rational alteration of 5’ mRNA folding free energy by introduction of synonymous mutations allows for predictable changes in pneumolysin (PLY) expression by Streptococcus pneumoniae without the need for chemical inducers or heterologous promoters. We created a panel of isogenic S. pneumoniae strains, differing only in synonymous (silent) mutations at the 5’ end of the PLY mRNA that are predicted to alter ΔG. Such manipulation allows rheostat-like control of PLY production and alters the cytotoxicity of whole S. pneumoniae on primary and immortalized human cells. These studies provide proof-of-principle for further investigation of mRNA ΔG manipulation as a tool in studies of bacterial pathogenesis. Public Library of Science 2015-03-23 /pmc/articles/PMC4370707/ /pubmed/25798590 http://dx.doi.org/10.1371/journal.pone.0119823 Text en © 2015 Amaral 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, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Amaral, Fábio E. Parker, Dane Randis, Tara M. Kulkarni, Ritwij Prince, Alice S. Shirasu-Hiza, Mimi M. Ratner, Adam J. Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae |
title | Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
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title_full | Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
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title_fullStr | Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
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title_full_unstemmed | Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
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title_short | Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae
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title_sort | rational manipulation of mrna folding free energy allows rheostat control of pneumolysin production by streptococcus pneumoniae |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370707/ https://www.ncbi.nlm.nih.gov/pubmed/25798590 http://dx.doi.org/10.1371/journal.pone.0119823 |
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