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Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production
Deoxynivalenol (DON) contamination of cereal grains caused by Fusarium head blight may be addressed by future RNA interference (RNAi)-based gene silencing approaches. However, utilizing these approaches will require a greater understanding of the principles that govern RNAi effectiveness in the path...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6116998/ https://www.ncbi.nlm.nih.gov/pubmed/30161200 http://dx.doi.org/10.1371/journal.pone.0202798 |
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author | Baldwin, Thomas Islamovic, Emir Klos, Kathy Schwartz, Paul Gillespie, James Hunter, Samuel Bregitzer, Phil |
author_facet | Baldwin, Thomas Islamovic, Emir Klos, Kathy Schwartz, Paul Gillespie, James Hunter, Samuel Bregitzer, Phil |
author_sort | Baldwin, Thomas |
collection | PubMed |
description | Deoxynivalenol (DON) contamination of cereal grains caused by Fusarium head blight may be addressed by future RNA interference (RNAi)-based gene silencing approaches. However, utilizing these approaches will require a greater understanding of the principles that govern RNAi effectiveness in the pathogen Fusarium graminearum. RNAi in higher eukaryotes, including fungi, involves processing double stranded RNA (dsRNA) into small interfering RNA (siRNA) that silence gene expression based on base pair complementarity. This study examined virulence, DON production, and the small RNA (sRNA) populations in response to RNAi-based silencing of TRI6, a transcription factor that positively regulates DON synthesis via control of TRI5 expression. Silencing was accomplished via the expression of transgenes encoding inverted repeats targeting various regions of TRI6 (RNAi vectors). Transgene expression was associated with novel, TRI6-specific siRNAs. For RNAi vectors targeting the majority of TRI6 sequence (~600 bp), a discontinuous, repeatable pattern was observed in which most siRNAs mapped to specific regions of TRI6. Targeting shorter regions (250–350 bp) did not alter the siRNA populations corresponding to that region of TRI6. No phased processing was observed. The 5' base of ~83% of siRNAs was uracil, consistent with DICER processing and ARGONAUTE binding preferences for siRNA. Mutant lines showed TRI6 siRNA-associated reductions of TRI5 expression on toxin inducing media and DON in infected wheat and barley spikes. Shorter RNAi vectors resulted in variable levels of silencing that were less than for the ~600 bp RNAi vector, with a 343 bp RNAi vector targeting the 5’ end of TRI6 having the best silencing efficiency. This work identifies efficient shorter region for silencing of TRI6 and describes the patterns of siRNA corresponding to those regions. |
format | Online Article Text |
id | pubmed-6116998 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-61169982018-09-16 Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production Baldwin, Thomas Islamovic, Emir Klos, Kathy Schwartz, Paul Gillespie, James Hunter, Samuel Bregitzer, Phil PLoS One Research Article Deoxynivalenol (DON) contamination of cereal grains caused by Fusarium head blight may be addressed by future RNA interference (RNAi)-based gene silencing approaches. However, utilizing these approaches will require a greater understanding of the principles that govern RNAi effectiveness in the pathogen Fusarium graminearum. RNAi in higher eukaryotes, including fungi, involves processing double stranded RNA (dsRNA) into small interfering RNA (siRNA) that silence gene expression based on base pair complementarity. This study examined virulence, DON production, and the small RNA (sRNA) populations in response to RNAi-based silencing of TRI6, a transcription factor that positively regulates DON synthesis via control of TRI5 expression. Silencing was accomplished via the expression of transgenes encoding inverted repeats targeting various regions of TRI6 (RNAi vectors). Transgene expression was associated with novel, TRI6-specific siRNAs. For RNAi vectors targeting the majority of TRI6 sequence (~600 bp), a discontinuous, repeatable pattern was observed in which most siRNAs mapped to specific regions of TRI6. Targeting shorter regions (250–350 bp) did not alter the siRNA populations corresponding to that region of TRI6. No phased processing was observed. The 5' base of ~83% of siRNAs was uracil, consistent with DICER processing and ARGONAUTE binding preferences for siRNA. Mutant lines showed TRI6 siRNA-associated reductions of TRI5 expression on toxin inducing media and DON in infected wheat and barley spikes. Shorter RNAi vectors resulted in variable levels of silencing that were less than for the ~600 bp RNAi vector, with a 343 bp RNAi vector targeting the 5’ end of TRI6 having the best silencing efficiency. This work identifies efficient shorter region for silencing of TRI6 and describes the patterns of siRNA corresponding to those regions. Public Library of Science 2018-08-30 /pmc/articles/PMC6116998/ /pubmed/30161200 http://dx.doi.org/10.1371/journal.pone.0202798 Text en https://creativecommons.org/publicdomain/zero/1.0/ This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 (https://creativecommons.org/publicdomain/zero/1.0/) public domain dedication. |
spellingShingle | Research Article Baldwin, Thomas Islamovic, Emir Klos, Kathy Schwartz, Paul Gillespie, James Hunter, Samuel Bregitzer, Phil Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title | Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title_full | Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title_fullStr | Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title_full_unstemmed | Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title_short | Silencing efficiency of dsRNA fragments targeting Fusarium graminearum TRI6 and patterns of small interfering RNA associated with reduced virulence and mycotoxin production |
title_sort | silencing efficiency of dsrna fragments targeting fusarium graminearum tri6 and patterns of small interfering rna associated with reduced virulence and mycotoxin production |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6116998/ https://www.ncbi.nlm.nih.gov/pubmed/30161200 http://dx.doi.org/10.1371/journal.pone.0202798 |
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