CRISPR mediated transactivation in the human disease vector Aedes aegypti

Michelle Bui, Elena Dalla Benetta, Yuemei Dong, Yunchong Zhao, Ting Yang, Ming Li, Igor A. Antoshechkin, Anna Buchman, Vanessa Bottino-Rojas, Anthony A. James, Michael W. Perry, George Dimopoulos, Omar S. Akbari

Research output: Contribution to journalArticlepeer-review


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.

Original languageEnglish (US)
Article numbere1010842
JournalPLoS pathogens
Issue number1
StatePublished - Jan 2023

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology
  • Virology
  • Parasitology
  • Microbiology
  • Immunology


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