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CRISPR mediated transactivation in the human disease vector Aedes aegypti
As a major insect vector of multiple arboviruses, Aedes aegypti poses a significant global health and economic burden. A number of genetic engineering tools have been exploited to understand its biology with the goal of reducing its impact. For example, current tools have focused on knocking-down RN...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9888728/ https://www.ncbi.nlm.nih.gov/pubmed/36656895 http://dx.doi.org/10.1371/journal.ppat.1010842 |
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author | Bui, Michelle Dalla Benetta, Elena Dong, Yuemei Zhao, Yunchong Yang, Ting Li, Ming Antoshechkin, Igor A. Buchman, Anna Bottino-Rojas, Vanessa James, Anthony A. Perry, Michael W. Dimopoulos, George Akbari, Omar S. |
author_facet | Bui, Michelle Dalla Benetta, Elena Dong, Yuemei Zhao, Yunchong Yang, Ting Li, Ming Antoshechkin, Igor A. Buchman, Anna Bottino-Rojas, Vanessa James, Anthony A. Perry, Michael W. Dimopoulos, George Akbari, Omar S. |
author_sort | Bui, Michelle |
collection | PubMed |
description | As a major insect vector of multiple arboviruses, Aedes aegypti poses a significant global health and economic burden. A number of genetic engineering tools have been exploited to understand its biology with the goal of reducing its impact. For example, current tools have focused on knocking-down RNA transcripts, inducing loss-of-function mutations, or expressing exogenous DNA. However, methods for transactivating endogenous genes have not been developed. To fill this void, here we developed a CRISPR activation (CRISPRa) system in Ae. aegypti to transactivate target gene expression. Gene expression is activated through pairing a catalytically-inactive (‘dead’) Cas9 (dCas9) with a highly-active tripartite activator, VP64-p65-Rta (VPR) and synthetic guide RNA (sgRNA) complementary to a user defined target-gene promoter region. As a proof of concept, we demonstrate that engineered Ae. aegypti mosquitoes harboring a binary CRISPRa system can be used to effectively overexpress two developmental genes, even-skipped (eve) and hedgehog (hh), resulting in observable morphological phenotypes. We also used this system to overexpress the positive transcriptional regulator of the Toll immune pathway known as AaRel1, which resulted in a significant suppression of dengue virus serotype 2 (DENV2) titers in the mosquito. This system provides a versatile tool for research pathways not previously possible in Ae. aegypti, such as programmed overexpression of endogenous genes, and may aid in gene characterization studies and the development of innovative vector control tools. |
format | Online Article Text |
id | pubmed-9888728 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-98887282023-02-01 CRISPR mediated transactivation in the human disease vector Aedes aegypti Bui, Michelle Dalla Benetta, Elena Dong, Yuemei Zhao, Yunchong Yang, Ting Li, Ming Antoshechkin, Igor A. Buchman, Anna Bottino-Rojas, Vanessa James, Anthony A. Perry, Michael W. Dimopoulos, George Akbari, Omar S. PLoS Pathog Research Article As a major insect vector of multiple arboviruses, Aedes aegypti poses a significant global health and economic burden. A number of genetic engineering tools have been exploited to understand its biology with the goal of reducing its impact. For example, current tools have focused on knocking-down RNA transcripts, inducing loss-of-function mutations, or expressing exogenous DNA. However, methods for transactivating endogenous genes have not been developed. To fill this void, here we developed a CRISPR activation (CRISPRa) system in Ae. aegypti to transactivate target gene expression. Gene expression is activated through pairing a catalytically-inactive (‘dead’) Cas9 (dCas9) with a highly-active tripartite activator, VP64-p65-Rta (VPR) and synthetic guide RNA (sgRNA) complementary to a user defined target-gene promoter region. As a proof of concept, we demonstrate that engineered Ae. aegypti mosquitoes harboring a binary CRISPRa system can be used to effectively overexpress two developmental genes, even-skipped (eve) and hedgehog (hh), resulting in observable morphological phenotypes. We also used this system to overexpress the positive transcriptional regulator of the Toll immune pathway known as AaRel1, which resulted in a significant suppression of dengue virus serotype 2 (DENV2) titers in the mosquito. This system provides a versatile tool for research pathways not previously possible in Ae. aegypti, such as programmed overexpression of endogenous genes, and may aid in gene characterization studies and the development of innovative vector control tools. Public Library of Science 2023-01-19 /pmc/articles/PMC9888728/ /pubmed/36656895 http://dx.doi.org/10.1371/journal.ppat.1010842 Text en © 2023 Bui et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Bui, Michelle Dalla Benetta, Elena Dong, Yuemei Zhao, Yunchong Yang, Ting Li, Ming Antoshechkin, Igor A. Buchman, Anna Bottino-Rojas, Vanessa James, Anthony A. Perry, Michael W. Dimopoulos, George Akbari, Omar S. CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title | CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title_full | CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title_fullStr | CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title_full_unstemmed | CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title_short | CRISPR mediated transactivation in the human disease vector Aedes aegypti |
title_sort | crispr mediated transactivation in the human disease vector aedes aegypti |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9888728/ https://www.ncbi.nlm.nih.gov/pubmed/36656895 http://dx.doi.org/10.1371/journal.ppat.1010842 |
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