A CRISPR–Cas9 gene drive targeting doublesex causes complete population suppression in caged Anopheles gambiae mosquitoes

• Published in: Nature biotechnology Vol. 36; no. 11; pp. 1062 - 1066
• Main Authors: Kyrou, Kyros, Hammond, Andrew M, Galizi, Roberto, Kranjc, Nace, Burt, Austin, Beaghton, Andrea K, Nolan, Tony, Crisanti, Andrea
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01-12-2018; Nature Publishing Group
• Subjects: 42/41; 42/44; 42/70; 631/1647/1511; 631/1647/1513; Agriculture; Alleles; Alternative splicing; Anopheles gambiae; Aquatic insects; Bioinformatics; Biomedical Engineering/Biotechnology; Biomedicine; Biotechnology; Conserved sequence; CRISPR; Culicidae; Disruption; Egg production; Females; Fertility; Life Sciences; Malaria; Mosquitoes; Phenotypes; Sterility; Transcription; Vector-borne diseases
• ISSN: 1087-0156; 1546-1696
• DOI: 10.1038/nbt.4245

Complete population collapse of malaria vector
Anopheles gambiae
in cages is achieved using a gene drive that targets
doublesex
.
In the human malaria vector
Anopheles gambiae
, the gene
doublesex
(
Agdsx
) encodes two alternatively spliced transcripts,
dsx-female
(
AgdsxF
) and
dsx-male
(
AgdsxM
), that control differentiation of the two sexes. The female transcript, unlike the male, contains an exon (exon 5) whose sequence is highly conserved in all
Anopheles
mosquitoes so far analyzed. We found that CRISPR–Cas9-targeted disruption of the intron 4–exon 5 boundary aimed at blocking the formation of functional AgdsxF did not affect male development or fertility, whereas females homozygous for the disrupted allele showed an intersex phenotype and complete sterility. A CRISPR–Cas9 gene drive construct targeting this same sequence spread rapidly in caged mosquitoes, reaching 100% prevalence within 7–11 generations while progressively reducing egg production to the point of total population collapse. Owing to functional constraint of the target sequence, no selection of alleles resistant to the gene drive occurred in these laboratory experiments. Cas9-resistant variants arose in each generation at the target site but did not block the spread of the drive.