scholarly journals Development and testing of a novel Killer-Rescue self-limiting gene drive system in Drosophila melanogaster

2019 ◽  
Author(s):  
Sophia H. Webster ◽  
Michael R. Vella ◽  
Maxwell J. Scott
Keyword(s):  
Drosophila Melanogaster ◽  
Core Promoter ◽  
Expression System ◽  
Drive System ◽  
Gene Drive ◽  
Drosophila Suzukii ◽  
Population Replacement ◽  
Gal4 Expression ◽  
Rescue System ◽  
Release Ratio

AbstractWe report the development and laboratory testing of a novel Killer-Rescue (K-R) self-limiting gene drive system in Drosophila melanogaster. This K-R system utilizes the well-characterized Gal4/UAS binary expression system and the Gal4 inhibitor, Gal80. Three killer (K) lines were tested; these used either an autoregulated UAS-Gal4 or UAS-Gal4 plus UAS-hid transgene. One universal rescue (R) line was used, UAS-Gal80, to inhibit Gal4 expression. The K lines are lethal and cause death in the absence of R. We show that Gal4 RNA levels are high in the absence of R. Death is possibly due to transcriptional squelching from high levels of Gal4. When R is present, Gal4 activation of Gal80 would lead to inhibition of Gal4 and prevent overexpression. With a single release ratio of 2:1 engineered K-R to wildtype, we find that K drives R through the population while the percent of wild type individuals decreases each generation. The choice of core promoter for a UAS-Gal4 construct strongly influences the K-R system. With the strong hsp70 core promoter, K was very effective but was quickly lost from the population. With the weaker DSCP core promoter, K persisted for longer allowing the frequency of individuals with at least one copy of R to increase to over 98%. This simple gene drive system could be readily adapted to other species such as mosquito disease vectors for driving anti-viral or anti-parasite genes.SignificanceHere we report the development and testing of a novel self-limiting gene drive system, Killer-Rescue, in Drosophila melanogaster. This system is composed of an auto-regulated Gal4 Killer (K) and a Gal4-activated Gal80 Rescue (R). Overexpression of Gal4 is lethal but in the presence of R, activation of Gal80 leads to much lower levels of Gal4 and rescue of lethality. We demonstrate that with a single 2:1 engineered to wildtype release, more than 98% of the population carry R after eight generations. We discuss how this Killer-Rescue system may be used for population replacement in a human health pest, Aedes aegypti, or for population suppression in an agricultural pest, Drosophila suzukii.

scholarly journals Development and testing of a novel killer–rescue self-limiting gene drive system in Drosophila melanogaster

2020 ◽  
Vol 287 (1925) ◽  
pp. 20192994 ◽  
Author(s):  
Sophia H. Webster ◽  
Michael R. Vella ◽  
Maxwell J. Scott
Keyword(s):  
Drosophila Melanogaster ◽  
Aedes Aegypti ◽  
Drive System ◽  
R Gene ◽  
Agricultural Pest ◽  
Gene Drive ◽  
Wild Type ◽  
Drosophila Suzukii ◽  
Population Replacement ◽  
Population Suppression

Here we report the development and testing of a novel self-limiting gene drive system, Killer–Rescue (K–R), in Drosophila melanogaster . This system is composed of an autoregulated Gal4 Killer (K) and a Gal4-activated Gal80 Rescue (R). Overexpression of Gal4 is lethal, but in the presence of R activation of Gal80 leads to much lower levels of Gal4 and rescue of lethality. We demonstrate that with a single 2 : 1 engineered to wild-type release, K drives R through the population and after nine generations, more than 98% of the population carry R and less than 2% of the population are wild-type flies. We discuss how this simple K–R gene drive system may be readily adapted for population replacement in a human health pest, Aedes aegypti , or for population suppression in an agricultural pest, Drosophila suzukii .

scholarly journals Molecular safeguarding of CRISPR gene drive experiments

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Jackson Champer ◽  
Joan Chung ◽  
Yoo Lim Lee ◽  
Chen Liu ◽  
Emily Yang ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Natural Population ◽  
Early Embryo ◽  
Drive System ◽  
Target Site ◽  
Gene Drive ◽  
Gene Drives ◽  
Synthetic Target

CRISPR-based homing gene drives have sparked both enthusiasm and deep concerns due to their potential for genetically altering entire species. This raises the question about our ability to prevent the unintended spread of such drives from the laboratory into a natural population. Here, we experimentally demonstrate the suitability of synthetic target site drives as well as split drives as flexible safeguarding strategies for gene drive experiments by showing that their performance closely resembles that of standard homing drives in Drosophila melanogaster. Using our split drive system, we further find that maternal deposition of both Cas9 and gRNA is required to form resistance alleles in the early embryo and that maternally-deposited Cas9 alone can power germline drive conversion in individuals that lack a genomic source of Cas9.

scholarly journals Genomic insertion locus and Cas9 expression in the germline affect CRISPR/Cas9-based gene drive performance in the yellow fever mosquito Aedes aegypti

2021 ◽  
Author(s):  
William R Reid ◽  
Jingyi Lin ◽  
Adeline E Williams ◽  
Rucsanda Juncu ◽  
Ken E Olson ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Yellow Fever ◽  
Drive System ◽  
Gene Drive ◽  
Yellow Fever Mosquito ◽  
Genomic Locus ◽  
Population Replacement ◽  
Novel Approach ◽  
Drive Systems ◽  
Insertion Locus

The yellow fever mosquito Aedes aegypti is a major vector of arthropod-borne viruses, including dengue, chikungunya, and Zika. A novel approach to mitigate arboviral infections is to generate mosquitoes refractory to infection by overexpressing antiviral effector molecules. Such an approach requires a mechanism to spread these antiviral effectors through a population, for example, by using CRISPR/Cas9-based gene drive systems. Here we report an autonomous single-component gene drive system in Ae. aegypti that is designed for persistent population replacement. Critical to the design of a single-locus autonomous gene drive is that the selected genomic locus be amenable to both gene drive and the appropriate expression of the antiviral effector. In our study, we took a reverse engineering approach to target two genomic loci ideal for the expression of antiviral effectors and further investigated the use of three promoters for Cas9 expression (nanos, β2-tubulin, or zpg) for the gene drive. We found that both promoter selection and genomic target site strongly influenced the efficiency of the drive, resulting in 100% inheritance in some crosses. We also observed the formation of inheritable gene drive blocking indels (GDBI) in the genomic locus with the highest levels of gene drive. Overall, our drive system forms a platform for the further testing of driving antipathogen effector genes through Ae. aegypti populations.

scholarly journals Gene Drives Across Engineered Fitness Valleys: Modeling a Design to Prevent Drive Spillover

2020 ◽  
Author(s):  
Frederik J.H. de Haas ◽  
Sarah P. Otto
Keyword(s):  
Local Population ◽  
Target Population ◽  
Drive System ◽  
High Threshold ◽  
Gene Drive ◽  
Time Model ◽  
Population Replacement ◽  
Low Threshold ◽  
Drive Systems ◽  
Gene Drives

1AbstractEngineered gene drive techniques for population replacement and/or suppression have potential for tackling complex challenges, including reducing the spread of diseases and invasive species. Unfortunately, the self-propelled behavior of drives can lead to the spread of transgenic elements beyond the target population, which is concerning. Gene drive systems with a low threshold frequency for invasion, such as homing-based gene drive systems, require initially few transgenic individuals to spread and are therefore easy to implement. However their ease of spread presents a double-edged sword; their low threshold makes these drives much more susceptible to spread outside of the target population (spillover). We model a proposed drive system that transitions in time from a low threshold drive system (homing-based gene drive) to a high threshold drive system (underdominance) using daisy chain technology. This combination leads to a spatially restricted drive strategy, while maintaining an attainable release threshold. We develop and analyze a discrete-time model as proof of concept and find that this technique effectively generates stable local population suppression, while preventing the spread of transgenic elements beyond the target population under biologically realistic parameters.

scholarly journals Synthetically engineered Medea gene drive system in the worldwide crop pest Drosophila suzukii

2018 ◽  
Vol 115 (18) ◽  
pp. 4725-4730 ◽  
Author(s):  
Anna Buchman ◽  
John M. Marshall ◽  
Dennis Ostrovski ◽  
Ting Yang ◽  
Omar S. Akbari
Keyword(s):  
Mendelian Inheritance ◽  
Drive System ◽  
Gene Drive ◽  
Fitness Costs ◽  
Drosophila Suzukii ◽  
Toxin Resistance ◽  
Crop Pest ◽  
Naturally Occurring ◽  
Drive Systems ◽  
Significant Disease

Synthetic gene drive systems possess enormous potential to replace, alter, or suppress wild populations of significant disease vectors and crop pests; however, their utility in diverse populations remains to be demonstrated. Here, we report the creation of a synthetic Medea gene drive system in a major worldwide crop pest, Drosophila suzukii. We demonstrate that this drive system, based on an engineered maternal “toxin” coupled with a linked embryonic “antidote,” is capable of biasing Mendelian inheritance rates with up to 100% efficiency. However, we find that drive resistance, resulting from naturally occurring genetic variation and associated fitness costs, can be selected for and hinder the spread of such a drive. Despite this, our results suggest that this gene drive could maintain itself at high frequencies in a wild population and spread to fixation if either its fitness costs or toxin resistance were reduced, providing a clear path forward for developing future such systems in this pest.

scholarly journals The mating rate of Drosophila suzukii reduction due to reproductive interference from Drosophila melanogaster

2020 ◽  
Author(s):  
Yongzhuo Chen ◽  
Min Zhang ◽  
Wei Hu ◽  
Jing Li ◽  
Pengcheng Liu ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Interspecific Competition ◽  
Population Size ◽  
Egg Laying ◽  
Significant Decline ◽  
Reproductive Interference ◽  
Drosophila Suzukii ◽  
Offspring Number ◽  
Successful Mating ◽  
Reproductive Activities

Abstract Background Drosophila suzukii is widely distributed. Research has revealed that the presence of Drosophila melanogaster can reduce the emergence and egg laying of D. suzukii. However, the reasons for these phenomena have not yet been reported. To investigate this issue, we sought to answer three questions: Can the presence of D. melanogaster reduce the longevity of D. suzukii? Does D. melanogaster dominate in larval interspecific competition with D. suzukii? Does reproductive interference occur between these species; i.e., do individuals of one species (e.g., D. suzukii) engage in reproductive activities with individuals of the other (e.g., D. melanogaster) such that the fitness of one or both species is reduced? Results The results showed that the adult offspring number of Drosophila suzukii was significantly reduced when this species was reared with Drosophila melanogaster. The larval interspecific competition had no significant effects on Drosophila suzukii longevity or population size. Surprisingly, Drosophila melanogaster imposed reproductive interference on males of Drosophila suzukii, which led to a significant decline in the rate of successful mating of the latter species. Conclusions The presence of Drosophila melanogaster causes the population size of Drosophila suzukii to decrease through reproductive interference, and the rate of successful mating in Drosophila suzukii is significantly reduced in the presence of Drosophila melanogaster.

scholarly journals One prophage WO gene rescues cytoplasmic incompatibility in Drosophila melanogaster

2018 ◽  
Author(s):  
J. Dylan Shropshire ◽  
Jungmin On ◽  
Emily M. Layton ◽  
Helen Zhou ◽  
Seth R. Bordenstein
Keyword(s):  
Drosophila Melanogaster ◽  
Vector Control ◽  
Genetic Basis ◽  
Cytoplasmic Incompatibility ◽  
Embryo Rescue ◽  
Field Trials ◽  
Current Control ◽  
Drive System ◽  
World Health ◽  
Fitness Advantage

AbstractWolbachia are maternally-inherited, intracellular bacteria at the forefront of vector control efforts to curb arbovirus transmission. In international field trials, the cytoplasmic incompatibility (CI) drive system of wMel Wolbachia is deployed to replace target vector populations, whereby a Wolbachia– induced modification of the sperm genome kills embryos. However, Wolbachia in the embryo rescue the sperm genome impairment, and therefore CI results in a strong fitness advantage for infected females that transmit the bacteria to offspring. The two genes responsible for the wMel-induced sperm modification of CI, cifA and cifB, were recently identified in the eukaryotic association module of prophage WO, but the genetic basis of rescue is unresolved. Here we use transgenic and cytological approaches to demonstrate that cifA independently rescues CI and nullifies embryonic death caused by wMel Wolbachia in Drosophila melanogaster. Discovery of cifA as the rescue gene and previously one of two CI induction genes establishes a new ‘Two-by-One’ model that underpins the genetic basis of CI. Results highlight the central role of prophage WO in shaping Wolbachia phenotypes that are significant to arthropod evolution and vector control.Significance StatementThe World Health Organization recommended pilot deployment of Wolbachia-infected mosquitoes to curb viral transmission to humans. Releases of mosquitoes are underway worldwide because Wolbachia can block replication of these pathogenic viruses and deterministically spread by a drive system termed cytoplasmic incompatibility (CI). Despite extensive research, the underlying genetic basis of CI remains only half-solved. We recently reported that two prophage WO genes recapitulate the modification component of CI in a released strain for vector control. Here we show that one of these genes underpins rescue of CI. Together, our results reveal the complete genetic basis of this selfish trait and pave the way for future studies exploring WO prophage genes as adjuncts or alternatives to current control efforts.

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