Fitness effects of CRISPR endonucleases in Drosophila melanogaster populations
- Cornell University, United States
- Peking University, China
Abstract
CRISPR/Cas9 provides a highly efficient and flexible genome editing technology with numerous potential applications ranging from gene therapy to population control. Some proposed applications involve the integration of CRISPR/Cas9 endonucleases into an organism's genome, which raises questions about potentially harmful effects to the transgenic individuals. One example for which this is particularly relevant are CRISPR-based gene drives conceived for the genetic alteration of entire populations. The performance of such drives can strongly depend on fitness costs experienced by drive carriers, yet relatively little is known about the magnitude and causes of these costs. Here, we assess the fitness effects of genomic CRISPR/Cas9 expression in Drosophila melanogaster cage populations by tracking allele frequencies of four different transgenic constructs that allow us to disentangle 'direct' fitness costs due to the integration, expression, and target-site activity of Cas9, from fitness costs due to potential off-target cleavage. Using a maximum likelihood framework, we find that a model with no direct fitness costs but moderate costs due to off-target effects fits our cage data best. Consistent with this, we do not observe fitness costs for a construct with Cas9HF1, a high-fidelity version of Cas9. We further demonstrate that using Cas9HF1 instead of standard Cas9 in a homing drive achieves similar drive conversion efficiency. These results suggest that gene drives should be designed with high-fidelity endonucleases and may have implications for other applications that involve genomic integration of CRISPR endonucleases.
Data availability
All data generated or analyzed during this study are included in the manuscript and supporting files.
Article and author information
Author details
Anna M Langmüller
Funding
National Institutes of Health (R21AI130635)
- Jackson Champer
- Andrew G Clark
- Philipp W Messer
National Institutes of Health (F32AI138476)
- Jackson Champer
National Institutes of Health (R01GM127418)
- Philipp W Messer
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: All flies with an active homing gene drive system were kept at the Sarkaria Arthropod Research Laboratory at Cornell University under Arthropod Containment Level 2 protocols in accordance with USDA APHIS standards. All safety standards were approved by the Cornell University Institutional Biosafety Committee.
Copyright
© 2022, Langmüller et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Fitness effects of CRISPR endonucleases in Drosophila melanogaster populations
eLife 11:e71809.
https://doi.org/10.7554/eLife.71809
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The majority of highly polymorphic genes are related to immune functions and with over 100 alleles within a population, genes of the major histocompatibility complex (MHC) are the most polymorphic loci in vertebrates. How such extraordinary polymorphism arose and is maintained is controversial. One possibility is heterozygote advantage (HA), which can in principle maintain any number of alleles, but biologically explicit models based on this mechanism have so far failed to reliably predict the coexistence of significantly more than ten alleles. We here present an eco-evolutionary model showing that evolution can result in the emergence and maintenance of more than 100 alleles under HA if the following two assumptions are fulfilled: first, pathogens are lethal in the absence of an appropriate immune defence; second, the effect of pathogens depends on host condition, with hosts in poorer condition being affected more strongly. Thus, our results show that HA can be a more potent force in explaining the extraordinary polymorphism found at MHC loci than currently recognized.
