scholarly journals Small-molecule control of super-Mendelian inheritance in gene drives

2019 ◽  
Author(s):  
Víctor López Del Amo ◽  
Brittany S. Leger ◽  
Kurt J. Cox ◽  
Shubhroz Gill ◽  
Alena L. Bishop ◽  
...  
Keyword(s):  
Small Molecule ◽  
Chemical Control ◽  
Control Strategies ◽  
Mendelian Inheritance ◽  
Drive System ◽  
Gene Drive ◽  
Crop Pests ◽  
Vector Borne Diseases ◽  
Vector Borne ◽  
Gene Drives

ABSTRACTBy surpassing the 50% inheritance limit of Mendel’s law of independent assortment, CRISPR-based gene drives have the potential to fight vector-borne diseases or suppress crop pests. However, contemporary gene drives could spread unchecked, posing safety concerns that limit their use in both laboratory and field settings. Current technologies also lack chemical control strategies, which could be applied in the field for dose, spatial and temporal control of gene drives. We describe in Drosophila the first gene-drive system controlled by an engineered Cas9 and a synthetic, orally-available small molecule.Graphical Abstract

scholarly journals Split-gene drive system provides flexible application for safe laboratory investigation and potential field deployment

2019 ◽  
Author(s):  
Víctor López Del Amo ◽  
Alena L. Bishop ◽  
Héctor M. Sánchez C. ◽  
Jared B. Bennett ◽  
Xuechun Feng ◽  
...  
Keyword(s):  
Mendelian Inheritance ◽  
Gene Drive ◽  
Island Conservation ◽  
Vector Borne Diseases ◽  
Split Gene ◽  
Mendelian Genetics ◽  
Vector Borne ◽  
Field Deployment ◽  
Flexible Application ◽  
Gene Drives

ABSTRACTCRISPR-based gene drives spread through populations bypassing the dictates of Mendelian genetics, offering a population-engineering tool for tackling vector-borne diseases, managing crop pests, and helping island conservation efforts; unfortunately, current technologies raise safety concerns for unintended gene propagation. Herein, we address this by splitting the two drive components, Cas9 and gRNAs, into separate alleles to form a novel trans-complementing split–gene-drive (tGD) and demonstrate its ability to promote super-Mendelian inheritance of the separate transgenes. This bi-component nature allows for individual transgene optimization and increases safety by restricting escape concerns to experimentation windows. We employ the tGD and a small– molecule-controlled version to investigate the biology of component inheritance and use our system to study the maternal effects on CRISPR inheritance, impaired homology on efficiency, and resistant allele formation. Lastly, mathematical modeling of tGD spread in a population shows potential advantages for improving current gene-drive technologies for field population modification.

scholarly journals Reducing resistance allele formation in CRISPR gene drive

2018 ◽  
Vol 115 (21) ◽  
pp. 5522-5527 ◽  
Author(s):  
Jackson Champer ◽  
Jingxian Liu ◽  
Suh Yeon Oh ◽  
Riona Reeves ◽  
Anisha Luthra ◽  
...  
Keyword(s):  
Natural Populations ◽  
Mendelian Inheritance ◽  
Gene Drive ◽  
Subsequent Formation ◽  
Vector Borne Diseases ◽  
Effective Strategies ◽  
Guide Rnas ◽  
Vector Borne ◽  
Resistance Rates ◽  
Gene Drives

CRISPR homing gene drives can convert heterozygous cells with one copy of the drive allele into homozygotes, thereby enabling super-Mendelian inheritance. Such a mechanism could be used, for example, to rapidly disseminate a genetic payload in a population, promising effective strategies for the control of vector-borne diseases. However, all CRISPR homing gene drives studied in insects thus far have produced significant quantities of resistance alleles that would limit their spread. In this study, we provide an experimental demonstration that multiplexing of guide RNAs can both significantly increase the drive conversion efficiency and reduce germline resistance rates of a CRISPR homing gene drive inDrosophila melanogaster. We further show that an autosomal drive can achieve drive conversion in the male germline, with no subsequent formation of resistance alleles in embryos through paternal carryover of Cas9. Finally, we find that thenanospromoter significantly lowers somatic Cas9 expression compared with thevasapromoter, suggesting thatnanosprovides a superior choice in drive strategies where gene disruption in somatic cells could have fitness costs. Comparison of drive parameters among the different constructs developed in this study and a previous study suggests that, while drive conversion and germline resistance rates are similar between different genomic targets, embryo resistance rates can vary significantly. Taken together, our results mark an important step toward developing effective gene drives capable of functioning in natural populations and provide several possible avenues for further control of resistance rates.

scholarly journals A CRISPR homing gene drive targeting a haplolethal gene removes resistance alleles and successfully spreads through a cage population

2020 ◽  
Vol 117 (39) ◽  
pp. 24377-24383 ◽  
Author(s):  
Jackson Champer ◽  
Emily Yang ◽  
Esther Lee ◽  
Jingxian Liu ◽  
Andrew G. Clark ◽  
...  
Keyword(s):  
Gene Drive ◽  
End Joining ◽  
Large Cage ◽  
Vector Borne Diseases ◽  
Guide Rnas ◽  
Genetic Modifications ◽  
New Strategy ◽  
Cage Population ◽  
Vector Borne ◽  
Gene Drives

Engineered gene drives are being explored as a new strategy in the fight against vector-borne diseases due to their potential for rapidly spreading genetic modifications through a population. However, CRISPR-based homing gene drives proposed for this purpose have faced a major obstacle in the formation of resistance alleles that prevent Cas9 cleavage. Here, we present a homing drive in Drosophila melanogaster that reduces the prevalence of resistance alleles below detectable levels by targeting a haplolethal gene with two guide RNAs (gRNAs) while also providing a rescue allele. Resistance alleles that form by end-joining repair typically disrupt the haplolethal target gene and are thus removed from the population because individuals that carry them are nonviable. We demonstrate that our drive is highly efficient, with 91% of the progeny of drive heterozygotes inheriting the drive allele and with no functional resistance alleles observed in the remainder. In a large cage experiment, the drive allele successfully spread to all individuals within a few generations. These results show that a haplolethal homing drive can provide an effective tool for targeted genetic modification of entire populations.

scholarly journals Modulating CRISPR gene drive activity through nucleocytoplasmic localization of Cas9 in S. cerevisiae

2018 ◽  
Author(s):  
Megan E. Goeckel ◽  
Erianna M. Basgall ◽  
Isabel C. Lewis ◽  
Samantha C. Goetting ◽  
Yao Yan ◽  
...  
Keyword(s):  
Nuclease Activity ◽  
Gene Drive ◽  
Wild Populations ◽  
Vector Borne Diseases ◽  
Vector Borne ◽  
Mendelian Laws ◽  
Artificial Gene ◽  
Nuclear Localization Sequences ◽  
Gene Drives

ABSTRACTThe bacterial CRISPR/Cas genome editing system has provided a major breakthrough in molecular biology. One use of this technology is within a nuclease-based gene drive. This type of system can install a genetic element within a population at unnatural rates. Combatting of vector-borne diseases carried by metazoans could benefit from a delivery system that bypasses traditional Mendelian laws of segregation. Recently, laboratory studies in fungi, insects, and even mice, have demonstrated successful propagation of CRISPR gene drives and the potential utility of this type of mechanism. However, current gene drives still face challenges including evolved resistance, containment, and the consequences of application in wild populations. In this study, we use an artificial gene drive system in budding yeast to explore mechanisms to modulate nuclease activity of Cas9 through its nucleocytoplasmic localization. We examine non-native nuclear localization sequences on Cas9 fusion proteins in vivo and demonstrate that appended signals can titrate gene drive activity and serve as a potential molecular safeguard.

scholarly journals A genetically encoded anti-CRISPR protein constrains gene drive spread and prevents population suppression

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Chrysanthi Taxiarchi ◽  
Andrea Beaghton ◽  
Nayomi Illansinhage Don ◽  
Kyros Kyrou ◽  
Matthew Gribble ◽  
...  
Keyword(s):  
Genetically Modified Organisms ◽  
Reproductive Capacity ◽  
Gene Drive ◽  
Vector Borne Diseases ◽  
Pathogen Effectors ◽  
Germline Expression ◽  
Vector Borne ◽  
Malaria Mosquitoes ◽  
Gene Drives ◽  
Population Suppression

AbstractCRISPR-based gene drives offer promising means to reduce the burden of pests and vector-borne diseases. These techniques consist of releasing genetically modified organisms carrying CRISPR-Cas nucleases designed to bias their inheritance and rapidly propagate desired modifications. Gene drives can be intended to reduce reproductive capacity of harmful insects or spread anti-pathogen effectors through wild populations, even when these confer fitness disadvantages. Technologies capable of halting the spread of gene drives may prove highly valuable in controlling, counteracting, and even reverting their effect on individual organisms as well as entire populations. Here we show engineering and testing of a genetic approach, based on the germline expression of a phage-derived anti-CRISPR protein (AcrIIA4), able to inactivate CRISPR-based gene drives and restore their inheritance to Mendelian rates in the malaria vector Anopheles gambiae. Modeling predictions and cage testing show that a single release of male mosquitoes carrying the AcrIIA4 protein can block the spread of a highly effective suppressive gene drive preventing population collapse of caged malaria mosquitoes.

scholarly journals Reducing resistance allele formation in CRISPR gene drives

2017 ◽  
Author(s):  
Jackson Champer ◽  
Jingxian Liu ◽  
Suh Yeon Oh ◽  
Riona Reeves ◽  
Anisha Luthra ◽  
...  
Keyword(s):  
Formation Rate ◽  
Natural Populations ◽  
Mendelian Inheritance ◽  
Resistance Allele ◽  
Subsequent Formation ◽  
Vector Borne Diseases ◽  
Male Germline ◽  
Vector Borne ◽  
Resistance Rates ◽  
Gene Drives

ABSTRACTCRISPR gene drives can efficiently convert heterozygous cells with one copy of the drive allele into homozygotes, thereby enabling super-Mendelian inheritance. This mechanism could be used, for example, to rapidly disseminate a genetic payload through a population, promising novel strategies for the control of vector-borne diseases. However, all CRISPR gene drives tested have produced significant quantities of resistance alleles that cannot be converted to drive alleles and would likely prevent these drives from spreading in a natural population. In this study, we assessed three strategies for reducing resistance allele formation. First, we directly compared drives with thenanosandvasapromoters, which showed that thevasadrive produced high levels of resistance alleles in somatic cells. This was not observed in thenanosdrive. Another strategy was the addition of a second gRNA to the drive, which both significantly increased the drive conversion efficiency and reduced the formation rate of resistance alleles. Finally, to minimize maternal carryover of Cas9, we assessed the performance of an autosomal drive acting in the male germline, and found no subsequent formation of resistance alleles in embryos. Our results mark a step toward developing effective gene drives capable of functioning in natural populations and provide several possible avenues for further reduction of resistance rates.

scholarly journals Resistance is futile: A CRISPR homing gene drive targeting a haplolethal gene

2019 ◽  
Author(s):  
Jackson Champer ◽  
Emily Yang ◽  
Yoo Lim Lee ◽  
Jingxian Liu ◽  
Andrew G. Clark ◽  
...  
Keyword(s):  
Target Gene ◽  
Gene Drive ◽  
End Joining ◽  
Large Cage ◽  
Vector Borne Diseases ◽  
Genetic Modifications ◽  
Vector Borne ◽  
Potential Strategy ◽  
Gene Drives ◽  
Population Suppression

ABSTRACTEngineered gene drives are being explored as a potential strategy for the control of vector-borne diseases due to their ability to rapidly spread genetic modifications through a population. While an effective CRISPR homing gene drive for population suppression has recently been demonstrated in mosquitoes, formation of resistance alleles that prevent Cas9 cleavage remains the major obstacle for drive strategies aiming at population modification, rather than elimination. Here, we present a homing drive in Drosophila melanogaster that reduces resistance allele formation below detectable levels by targeting a haplolethal gene with two gRNAs while also providing a rescue allele. This is because any resistance alleles that form by end-joining repair will typically disrupt the haplolethal target gene, rendering the individuals carrying them nonviable. We demonstrate that our drive is highly efficient, with 91% of the progeny of drive heterozygotes inheriting the drive allele and with no resistance alleles observed in the remainder. In a large cage experiment, the drive allele successfully spread to all individuals. These results show that a haplolethal homing drive can be a highly effective tool for population modification.

scholarly journals Wolbachia: endosymbiont of onchocercid nematodes and their vectors

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Ranju Ravindran Santhakumari Manoj ◽  
Maria Stefania Latrofa ◽  
Sara Epis ◽  
Domenico Otranto
Keyword(s):  
Control Strategies ◽  
Molecular Data ◽  
Scientific Data ◽  
Growth And Survival ◽  
Vector Borne Diseases ◽  
Wide Range ◽  
Veterinary Importance ◽  
Wolbachia Endosymbiont ◽  
Vector Borne ◽  
And Control

Abstract Background Wolbachia is an obligate intracellular maternally transmitted, gram-negative bacterium which forms a spectrum of endosymbiotic relationships from parasitism to obligatory mutualism in a wide range of arthropods and onchocercid nematodes, respectively. In arthropods Wolbachia produces reproductive manipulations such as male killing, feminization, parthenogenesis and cytoplasmic incompatibility for its propagation and provides an additional fitness benefit for the host to protect against pathogens, whilst in onchocercid nematodes, apart from the mutual metabolic dependence, this bacterium is involved in moulting, embryogenesis, growth and survival of the host. Methods This review details the molecular data of Wolbachia and its effect on host biology, immunity, ecology and evolution, reproduction, endosymbiont-based treatment and control strategies exploited for filariasis. Relevant peer-reviewed scientic papers available in various authenticated scientific data bases were considered while writing the review. Conclusions The information presented provides an overview on Wolbachia biology and its use in the control and/or treatment of vectors, onchocercid nematodes and viral diseases of medical and veterinary importance. This offers the development of new approaches for the control of a variety of vector-borne diseases. Graphic Abstract

Towards CRISPR/Cas9-based gene drive in the diamondback moth Plutella xylostella

2021 ◽  
Author(s):  
Xuejiao Xu ◽  
Tim Harvey-Samuel ◽  
Hamid Anees Siddiqui ◽  
Joshua Ang ◽  
Michelle E Anderson ◽  
...  
Keyword(s):  
Genetic Control ◽  
Plutella Xylostella ◽  
Control Strategies ◽  
Diamondback Moth ◽  
Regulatory Elements ◽  
Strategy Development ◽  
Gene Drive ◽  
Global Agriculture ◽  
High Level ◽  
Gene Drives

Promising to provide powerful genetic control tools, gene drives have been constructed in multiple dipterans, yeast and mice, for the purposes of population elimination or modification. However, it remains unclear whether these techniques can be applied to lepidopterans. Here, we used endogenous regulatory elements to drive Cas9 and sgRNA expression in the diamondback moth, (Plutella xylostella), and test the first split-drive system in a lepidopteran. The diamondback moth is an economically important global agriculture pest of cruciferous crops and has developed severe resistance to various insecticides, making it a prime candidate for such novel control strategy development. A very high level of somatic editing was observed in Cas9/sgRNA transheterozygotes, although no significant homing was revealed in the subsequent generation. Although heritable, Cas9-medated germline cleavage, as well as maternal and paternal Cas9 deposition was observed, rates were far lower than for somatic cleavage events, indicating robust somatic but limited germline activity of Cas9/sgRNA under the control of selected regulatory elements. Our results provide valuable experience, paving the way for future construction of gene drive-based genetic control strategies in DBM or other lepidopterans.

Sign in / Sign up

Export Citation Format

Share Document