Intron-derived small RNAs for silencing viral RNAs in mosquito cells.
Aedes aegypti and Ae. albopictus are the main vectors of mosquito-borne viruses of medical and veterinary significance. Many of these viruses have RNA genomes. Exogenously provided, e.g. transgene encoded, small RNAs could be used to inhibit virus replication, breaking the transmission cycle. We tes...
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ftdoajarticles:oai:doaj.org/article:ff86f9512b254a0781e63007c15c834c 2023-06-11T04:09:45+02:00 Intron-derived small RNAs for silencing viral RNAs in mosquito cells. Priscilla Y L Tng Leonela Z Carabajal Paladino Michelle A E Anderson Zach N Adelman Rennos Fragkoudis Rob Noad Luke Alphey 2022-06-01T00:00:00Z https://doi.org/10.1371/journal.pntd.0010548 https://doaj.org/article/ff86f9512b254a0781e63007c15c834c EN eng Public Library of Science (PLoS) https://doi.org/10.1371/journal.pntd.0010548 https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0010548 https://doaj.org/article/ff86f9512b254a0781e63007c15c834c PLoS Neglected Tropical Diseases, Vol 16, Iss 6, p e0010548 (2022) Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 article 2022 ftdoajarticles https://doi.org/10.1371/journal.pntd.0010548 2023-04-23T00:32:01Z Aedes aegypti and Ae. albopictus are the main vectors of mosquito-borne viruses of medical and veterinary significance. Many of these viruses have RNA genomes. Exogenously provided, e.g. transgene encoded, small RNAs could be used to inhibit virus replication, breaking the transmission cycle. We tested, in Ae. aegypti and Ae. albopictus cell lines, reporter-based strategies for assessing the ability of two types of small RNAs to inhibit a chikungunya virus (CHIKV) derived target. Both types of small RNAs use a Drosophila melanogaster pre-miRNA-1 based hairpin for their expression, either with perfect base-pairing in the stem region (shRNA-like) or containing two mismatches (miRNA-like). The pre-miRNA-1 stem loop structure was encoded within an intron; this allows co-expression of one or more proteins, e.g. a fluorescent protein marker tracking the temporal and spatial expression of the small RNAs in vivo. Three reporter-based systems were used to assess the relative silencing efficiency of ten shRNA-like siRNAs and corresponding miRNA-like designs. Two systems used a luciferase reporter RNA with CHIKV RNA inserted either in the coding sequence or within the 3' UTR. A third reporter used a CHIKV derived split replication system. All three reporters demonstrated that while silencing could be achieved with both miRNA-like and shRNA-like designs, the latter were substantially more effective. Dcr-2 was required for the shRNA-like siRNAs as demonstrated by loss of inhibition of the reporters in Dcr-2 deficient cell lines. These positive results in cell culture are encouraging for the potential use of this pre-miRNA-1-based system in transgenic mosquitoes. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic PLOS Neglected Tropical Diseases 16 6 e0010548 |
institution |
Open Polar |
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Directory of Open Access Journals: DOAJ Articles |
op_collection_id |
ftdoajarticles |
language |
English |
topic |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
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Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 Priscilla Y L Tng Leonela Z Carabajal Paladino Michelle A E Anderson Zach N Adelman Rennos Fragkoudis Rob Noad Luke Alphey Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
description |
Aedes aegypti and Ae. albopictus are the main vectors of mosquito-borne viruses of medical and veterinary significance. Many of these viruses have RNA genomes. Exogenously provided, e.g. transgene encoded, small RNAs could be used to inhibit virus replication, breaking the transmission cycle. We tested, in Ae. aegypti and Ae. albopictus cell lines, reporter-based strategies for assessing the ability of two types of small RNAs to inhibit a chikungunya virus (CHIKV) derived target. Both types of small RNAs use a Drosophila melanogaster pre-miRNA-1 based hairpin for their expression, either with perfect base-pairing in the stem region (shRNA-like) or containing two mismatches (miRNA-like). The pre-miRNA-1 stem loop structure was encoded within an intron; this allows co-expression of one or more proteins, e.g. a fluorescent protein marker tracking the temporal and spatial expression of the small RNAs in vivo. Three reporter-based systems were used to assess the relative silencing efficiency of ten shRNA-like siRNAs and corresponding miRNA-like designs. Two systems used a luciferase reporter RNA with CHIKV RNA inserted either in the coding sequence or within the 3' UTR. A third reporter used a CHIKV derived split replication system. All three reporters demonstrated that while silencing could be achieved with both miRNA-like and shRNA-like designs, the latter were substantially more effective. Dcr-2 was required for the shRNA-like siRNAs as demonstrated by loss of inhibition of the reporters in Dcr-2 deficient cell lines. These positive results in cell culture are encouraging for the potential use of this pre-miRNA-1-based system in transgenic mosquitoes. |
format |
Article in Journal/Newspaper |
author |
Priscilla Y L Tng Leonela Z Carabajal Paladino Michelle A E Anderson Zach N Adelman Rennos Fragkoudis Rob Noad Luke Alphey |
author_facet |
Priscilla Y L Tng Leonela Z Carabajal Paladino Michelle A E Anderson Zach N Adelman Rennos Fragkoudis Rob Noad Luke Alphey |
author_sort |
Priscilla Y L Tng |
title |
Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
title_short |
Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
title_full |
Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
title_fullStr |
Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
title_full_unstemmed |
Intron-derived small RNAs for silencing viral RNAs in mosquito cells. |
title_sort |
intron-derived small rnas for silencing viral rnas in mosquito cells. |
publisher |
Public Library of Science (PLoS) |
publishDate |
2022 |
url |
https://doi.org/10.1371/journal.pntd.0010548 https://doaj.org/article/ff86f9512b254a0781e63007c15c834c |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
PLoS Neglected Tropical Diseases, Vol 16, Iss 6, p e0010548 (2022) |
op_relation |
https://doi.org/10.1371/journal.pntd.0010548 https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0010548 https://doaj.org/article/ff86f9512b254a0781e63007c15c834c |
op_doi |
https://doi.org/10.1371/journal.pntd.0010548 |
container_title |
PLOS Neglected Tropical Diseases |
container_volume |
16 |
container_issue |
6 |
container_start_page |
e0010548 |
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1768383740547956736 |