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Functional intron-derived miRNAs and host-gene expression in plants

BACKGROUND: Recently, putative pre-miRNAs locations have been identified in the introns of plant genes, raising the question whether such genes can show a dual functionality by having both correct maturation of the host gene pre-mRNA and maturation of the miRNAs from the intron. Here, we demonstrate...

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Autores principales: Shapulatov, Umidjon, van Hoogdalem, Mark, Schreuder, Marielle, Bouwmeester, Harro, Abdurakhmonov, Ibrokhim Y., van der Krol, Alexander R.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151947/
https://www.ncbi.nlm.nih.gov/pubmed/30258486
http://dx.doi.org/10.1186/s13007-018-0351-2
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author Shapulatov, Umidjon
van Hoogdalem, Mark
Schreuder, Marielle
Bouwmeester, Harro
Abdurakhmonov, Ibrokhim Y.
van der Krol, Alexander R.
author_facet Shapulatov, Umidjon
van Hoogdalem, Mark
Schreuder, Marielle
Bouwmeester, Harro
Abdurakhmonov, Ibrokhim Y.
van der Krol, Alexander R.
author_sort Shapulatov, Umidjon
collection PubMed
description BACKGROUND: Recently, putative pre-miRNAs locations have been identified in the introns of plant genes, raising the question whether such genes can show a dual functionality by having both correct maturation of the host gene pre-mRNA and maturation of the miRNAs from the intron. Here, we demonstrated that such dual functionality is indeed possible, using as host gene the firefly luciferase gene with intron (ffgLUC), and different artificial intronic miRNAs (aimiRNA) placed within the intron of ffgLUC. RESULTS: The miRNAs were based on the structure of the natural miR319a. Luciferase (LUC) activity in planta was used to evaluate a correct splicing of the ffgLUC mRNA. Different target sequences were inserted into the aimiRNA to monitor efficiency of silencing of different target mRNAs. After adjusting the insertion cloning strategy, the ffgLUC(aimiR-319a) gene showed dual functionality with correct splicing of ffgLUC and efficient silencing of TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR1 transcription factor genes targeted in-trans by aimiR-319a or targeting the transgene ffLUC in-cis by an aimiR-LUC. Silencing of endogenous target genes by aimiRNA or amiRNA is efficient both in transient assays and stable transformants. A behave as strong phenotype the PHYTOCHROME B (PHYB) gene was also targeted by ffgLUC(aimiR-PHYB). The lack of silencing of the PHYB target was most likely due to an insensitive target site within the PHYB mRNA which can potentially form a double stranded stem structure. CONCLUSION: The combination of an overexpression construct with an artificial intronic microRNA allows for a simultaneous dual function in plants. The concept therefore adds new options to engineering of plant traits that require multiple gene manipulations. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s13007-018-0351-2) contains supplementary material, which is available to authorized users.
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spelling pubmed-61519472018-09-26 Functional intron-derived miRNAs and host-gene expression in plants Shapulatov, Umidjon van Hoogdalem, Mark Schreuder, Marielle Bouwmeester, Harro Abdurakhmonov, Ibrokhim Y. van der Krol, Alexander R. Plant Methods Research BACKGROUND: Recently, putative pre-miRNAs locations have been identified in the introns of plant genes, raising the question whether such genes can show a dual functionality by having both correct maturation of the host gene pre-mRNA and maturation of the miRNAs from the intron. Here, we demonstrated that such dual functionality is indeed possible, using as host gene the firefly luciferase gene with intron (ffgLUC), and different artificial intronic miRNAs (aimiRNA) placed within the intron of ffgLUC. RESULTS: The miRNAs were based on the structure of the natural miR319a. Luciferase (LUC) activity in planta was used to evaluate a correct splicing of the ffgLUC mRNA. Different target sequences were inserted into the aimiRNA to monitor efficiency of silencing of different target mRNAs. After adjusting the insertion cloning strategy, the ffgLUC(aimiR-319a) gene showed dual functionality with correct splicing of ffgLUC and efficient silencing of TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR1 transcription factor genes targeted in-trans by aimiR-319a or targeting the transgene ffLUC in-cis by an aimiR-LUC. Silencing of endogenous target genes by aimiRNA or amiRNA is efficient both in transient assays and stable transformants. A behave as strong phenotype the PHYTOCHROME B (PHYB) gene was also targeted by ffgLUC(aimiR-PHYB). The lack of silencing of the PHYB target was most likely due to an insensitive target site within the PHYB mRNA which can potentially form a double stranded stem structure. CONCLUSION: The combination of an overexpression construct with an artificial intronic microRNA allows for a simultaneous dual function in plants. The concept therefore adds new options to engineering of plant traits that require multiple gene manipulations. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s13007-018-0351-2) contains supplementary material, which is available to authorized users. BioMed Central 2018-09-24 /pmc/articles/PMC6151947/ /pubmed/30258486 http://dx.doi.org/10.1186/s13007-018-0351-2 Text en © The Author(s) 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Shapulatov, Umidjon
van Hoogdalem, Mark
Schreuder, Marielle
Bouwmeester, Harro
Abdurakhmonov, Ibrokhim Y.
van der Krol, Alexander R.
Functional intron-derived miRNAs and host-gene expression in plants
title Functional intron-derived miRNAs and host-gene expression in plants
title_full Functional intron-derived miRNAs and host-gene expression in plants
title_fullStr Functional intron-derived miRNAs and host-gene expression in plants
title_full_unstemmed Functional intron-derived miRNAs and host-gene expression in plants
title_short Functional intron-derived miRNAs and host-gene expression in plants
title_sort functional intron-derived mirnas and host-gene expression in plants
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151947/
https://www.ncbi.nlm.nih.gov/pubmed/30258486
http://dx.doi.org/10.1186/s13007-018-0351-2
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