Biology subjects

Champer, J.

Publications and source records attributed to Champer, J..

At least 55 records · Page 3Linked to original sources

bioRxiv2023

A framework for identifying fertility gene targets for mammalian pest control

O_LIFertility-targeted gene drives have been proposed as an ethical genetic approach for managing wild populations of vertebrate pests for public health and conservation benefit. C_LIO_LIThis manuscript introduces a framework to identify and evaluate target gene suitability based on biological gene function, gene expression, and results from mouse knockout models. C_LIO_LIThis framework identified 16 genes essential for male fertility and 12 genes important for female fertility that may be feasible targets for mammalian gene drives and other non-drive genetic pest control technology. Further, a comparative genomics analysis demonstrates the conservation of the identified genes across several globally significant invasive mammals. C_LIO_LIIn addition to providing important considerations for identifying candidate genes, our framework and the genes identified in this study may have utility in developing additional pest control tools such as wildlife contraceptives. C_LI

genetics↗

bioRxiv2023

Assessment of distant-site rescue elements for CRISPR toxin-antidote gene drives

New types of gene drives promise to provide increased flexibility, offering many options for confined modification or suppression of target populations. Among the most promising are CRISPR toxin-antidote gene drives, which disrupt essential wild-type genes by targeting them with Cas9/gRNA, resulting in their removal. This increases the frequency of the drive in the population. All these drives, plus homing modification rescue drives, rely on having an effective rescue element, which consists of a recoded version of the target gene. This rescue element can be at the same site as the target gene, which maximizes the chance of efficient rescue, or at a distant site, which allows some other useful options, such as easily disrupting another essential gene or achieving greater confinement. Previously, we developed a homing rescue drive targeting a haplolethal gene and a toxin-antidote drive targeting an essential but haplosufficient gene. These successful drives had functional rescue elements but suboptimal drive efficiency. Here, we attempted to construct new toxin-antidote drives targeting these genes with a distantsite configuration from three different loci. We found that use of additional gRNAs increased cut rates to nearly 100%. However, all distant-site rescue elements failed for both haplolethal and haplosufficient target genes. Furthermore, one rescue element with a minimally recoded rescue element was used as a template for homology-directed repair for the target gene on a different chromosomal arm, resulting in the formation of functional resistance alleles at high frequency. Together, these results can inform the design of future CRISPR-based toxin-antidote gene drives.

genetics↗

bioRxiv2022

Performance characteristics allow for confinement of a CRISPR toxin-antidote gene drive designed for population suppression in a reaction-diffusion model

Gene drives alleles that can bias their own inheritance are a promising way to engineer populations for control of disease vectors, invasive species, and agricultural pests. Recent advancements in the field have yielded successful examples of powerful suppression type drives and confined modification type drives, but developing confined suppression drives has proven more difficult. This is because the necessary power for strong suppression is often incompatible with the characteristics needed for drive confinement. However, one type of CRISPR toxin-antidote drive may be strong enough and confined, the TADE (Toxin-Antidote Dominant Embryo) suppression drive. By disrupting a haplolethal target gene and a haplosufficient female fertility gene, this drive quickly eliminates wild-type alleles and eventually induces population suppression. It has been shown to perform effectively in panmictic populations. However, confinement in spatial scenarios may be substantially different. Here, we use a reaction-diffusion model to assess the performance of TADE suppression drive in continuous space. We measure the drive wave advance speed while varying several performance parameters and find that moderate fitness costs or embryo cutting (from maternally deposited nuclease) can eliminate the drives ability to form a wave of advance. We assess the release size required for the drive to propagate, and finally, we investigate migration corridor scenarios. Depending on the corridor size and dispersal, it is often possible for the drive to suppress one population and then persist in the corridor without invading the second population. This prevents re-invasion by wild-type, which may be a particularly desirable outcome in some scenarios. Thus, even imperfect variants of TADE suppression drive may be excellent candidates for confined population suppression.

genetics↗

bioRxiv2022

Making waves: Comparative analysis of gene drive spread characteristics in a continuous space model

With their ability to rapidly increase in frequency, gene drives can be used to modify or suppress target populations after an initial release of drive-containing individuals. Recent advances in this field have revealed many possibilities for different types of drives, and several of these have been realized in experimental demonstrations. These drives all have unique advantages and disadvantages related to their ease of construction, confinement, and capacity to act as a modification or suppression system. While many properties of these drives have been explored in modelling studies, assessment of these drives in continuous space environments has been limited, often focusing on outcomes rather than fundamental properties. Here, we conduct a comparative analysis of many different gene drive types that have the capacity to form a wave of advance against wild-type alleles in one-dimensional continuous space. We evaluate the drive wave speed as a function of drive performance and ecological parameters, which reveals substantial differences between drive performance in panmictic versus spatial environments. In particular, we find that suppression drive waves are uniquely vulnerable to fitness costs and undesired CRISPR cleavage activity that can form resistance alleles in embryos by maternal deposition. Some drives, though, retain robust characteristics even with widely varying performance characteristics. To gain a better understanding of drive waves, we compare panmictic performance of drives across the full range of drive frequencies. We find that rates of wild-type allele removal in panmictic setting is correlated with drive wave speed, though this is also affected by a range of other factors. Overall, our results provide a useful resource for understanding the performance of drives in continuous spatial environments, which may be most representative of potential drive deployment in many relevant scenarios.

genetics↗

bioRxiv2022

Simulations reveal high efficiency and confinement of a population suppression CRISPR toxin-antidote gene drive

Though engineered gene drives hold great promise for spreading through and eventually suppressing populations of disease vectors or invasive species, complications such as resistance alleles and spatial population structure can prevent their success. Additionally, most forms of suppression drives, such as homing drives or driving Y chromosomes, will generally spread uncontrollably between populations with even small levels of migration. The previously proposed CRISPR-based toxin-antidote system called TADE suppression drive could potentially address the issue of confinement and resistance alleles. However, it is a relatively weak form of drive compared to homing drives, which might make it particularly vulnerable to spatial population structure. In this study, we investigate TADE suppression drive using individual-based simulations in continuous space. We find that the drive is actually more confined in continuous space than in panmictic populations, even in its most efficient form with a low cleavage rate in embryos from maternally deposited Cas9. Furthermore, the drive performed well in continuous space scenarios if the initial release requirements were met, suppressing the populations in a timely manner without being severely affected by chasing, a phenomenon in which wild-type individuals avoid the drive by recolonizing empty areas. At higher embryo cut rates, the drive loses its ability to propagate on its own, but a single, widespread release can often still induce rapid population collapse. Thus, if TADE suppression gene drives can be successfully constructed, they may play an important role in control of disease vectors and invasive species when stringent confinement to target populations is desired.

evolutionary biology↗

bioRxiv2022

Harnessing Wolbachia cytoplasmic incompatibility alleles for confined gene drive: a modeling study

Wolbachia is a maternally-inherited bacteria, which can spread rapidly in populations by manipulating reproduction. CifA and CifB are genes found in Wolbachia phage that are responsible for cytoplasmic incompatibility, the most common type of Wolbachia reproductive interference. In this phenomenon, no viable offspring are produced when a male with both CifA and CifB (or just CifB in an alternative mechanism) mates with a female lacking CifA. Utilizing this feature, we propose new types of toxin-antidote gene drives that can be constructed with these genes. By using both mathematical and simulation models, we found that a drive containing CifA and CifB together create a confined drive with a moderate to high introduction threshold. When introduced separately, they act as a self-limiting drive. We observed that the performance of these drives is substantially influenced by various ecological parameters and drive characteristics. Extending our models to continuous space, we found that the drive individual release distribution has a critical impact on drive persistence. Our results suggest that these new types of drives based on Wolbachia transgenes are safe and flexible candidates for genetic modification of populations.

genetics↗

bioRxiv2022

Adversarial interspecies relationships facilitate population suppression by gene drive in spatially explicit models

Suppression gene drives are designed to bias their inheritance and increase in frequency in a population, disrupting an essential gene in the process. When the frequency is high enough, the population will be unable to reproduce above the replacement level and could be eliminated. CRISPR suppression drives based on the homing mechanism have already seen success in the laboratory, particularly in malaria mosquitoes. However, several models predict that the use of these drives in realistic populations with spatial structure may not achieve complete success. This is due to the ability of wild-type individuals to escape the drive and reach empty areas with reduced competition, allowing them to achieve high reproductive success and leading to extinction-recolonization cycles across the landscape. Here, we extend our continuous space gene drive framework to include two competing species or predator-prey species pairs. We find that in both general and mosquito-specific models, the presence of a competing species or predator can greatly facilitate drive-based suppression, even for drives with modest efficiency. However, the presence of a competing species also substantially increases the frequency of outcomes in which the drive is lost before suppression is achieved. These results are robust in models with seasonal population fluctuations that mosquito populations often experience. We also found that suppression can be somewhat more difficult if targeting a predator with strong predator-prey interactions. Our results illustrate the difficulty of predicting outcomes of interventions that could substantially affect populations of interacting species in complex ecosystems. However, our results are also potentially promising for the prospects of less powerful suppression gene drives for achieving successful elimination of target mosquito and other pest populations.

ecology↗

bioRxiv2022

Finding the strongest gene drive: Simulations reveal unexpected performance differences between Anopheles homing suppression drive candidates

Recent experiments have produced several Anopheles gambiae homing gene drives that disrupt female fertility genes, thereby eventually inducing population collapse. Such drives may be highly effective tools to combat malaria. One such homing drive, based on the zpg promoter driving CRISPR/Cas9, was able to eliminate a cage population of mosquitoes. A second version, purportedly improved upon the first by incorporating an X-shredder element (which biases inheritance towards male offspring), was similarly successful. Here, we re-analyze the data of each of these gene drives and suggest an alternative interpretation of their performance. We assess each suppression drive within an individual-based simulation framework that models mosquito population dynamics in continuous space. We find that the combined homing/X-shredder drive is actually less effective at population suppression within the context of our mosquito population model. In particular, the combined drive often fails to completely suppress the population, instead resulting in an unstable equilibrium between drive and wild-type alleles. By contrast, otherwise similar drives based on the nos promoter may prove to be more promising candidates for future development due to potentially superior performance.

genetics↗

bioRxiv2021

Modeling homing suppression gene drive in haplodiploid organisms

Gene drives have shown great promise for suppression of pest populations. These engineered alleles can function by a variety of mechanisms, but the most common is the CRISPR homing drive, which converts wild-type alleles to drive alleles in the germline of heterozygotes. Some potential target species are haplodiploid, in which males develop from unfertilized eggs and thus have only one copy of each chromosome. This prevents drive conversion, a substantial disadvantage compared to diploids where drive conversion can take place in both sexes. Here, we study homing suppression gene drives in haplodiploids and find that a drive targeting a female fertility gene could still be successful. However, such drives are less powerful than in diploids and suffer more from functional resistance alleles. They are substantially more vulnerable to high resistance allele formation in the embryo due to maternally deposited Cas9 and gRNA and also to somatic cleavage activity. Examining spatial models where organisms move over a continuous landscape, we find that haplodiploid suppression drives surprisingly perform nearly as well as in diploids, possibly due to their ability to spread further before inducing strong suppression. Together, these results indicate that gene drive can potentially be used to effectively suppress haplodiploid populations.

evolutionary biology↗

bioRxiv2021

Experimental demonstration of tethered gene drive systems for confined population modification or suppression

Homing gene drives hold great promise for the genetic control of natural populations. However, current homing systems are capable of spreading uncontrollably between populations connected by even marginal levels of migration. This could represent a substantial sociopolitical barrier to the testing or deployment of such drives and may generally be undesirable when the objective is only local population control, such as suppression of an invasive species outside of its native range. Tethered drive systems, in which a locally confined gene drive provides the CRISPR nuclease needed for a homing drive, could provide a solution to this problem, offering the power of a homing drive and confinement of the supporting drive. Here, we demonstrate the engineering of a tethered drive system in Drosophila, using a regionally confined CRISPR Toxin-Antidote Recessive Embryo (TARE) drive to support modification and suppression homing drives. Each drive was able to bias inheritance in its favor, and the TARE drive was shown to spread only when released above a threshold frequency in experimental cage populations. After the TARE drive had established in the population, it facilitated the spread of a subsequently released split homing modification drive (to all individuals in the cage) and of a homing suppression drive (to its equilibrium frequency). Our results show that the tethered drive strategy is a viable and easily engineered option for providing confinement of homing drives to target populations.

genetics↗

bioRxiv2021

A homing suppression gene drive with multiplexed gRNAs maintains high drive conversion efficiency and avoids functional resistance alleles

Gene drives are engineered alleles that can bias inheritance in their favor, allowing them to spread throughout a population. They could potentially be used to modify or suppress pest populations, such as mosquitoes that spread diseases. CRISPR/Cas9 homing drives, which copy themselves by homology-directed repair in drive/wild-type heterozygotes, are a powerful form of gene drive, but they are vulnerable to resistance alleles that preserve the function of their target gene. Such resistance alleles can prevent successful population suppression. Here, we constructed a homing suppression drive in Drosophila melanogaster that utilized multiplexed gRNAs to inhibit the formation of functional resistance alleles in its female fertility target gene. The selected gRNA target sites were close together, preventing reduction in drive conversion efficiency. The construct reached a moderate equilibrium frequency in cage populations without apparent formation of resistance alleles. However, a moderate fitness cost prevented elimination of the cage population, showing the importance of using highly efficient drives in a suppression strategy, even if resistance can be addressed. Nevertheless, our results experimentally demonstrate the viability of the multiplexed gRNAs strategy in homing suppression gene drives.

genetics↗

bioRxiv2021

Fitness effects of CRISPR endonucleases in Drosophila melanogaster populations

CRISPR/Cas9 systems provide a highly efficient and flexible genome editing technology with numerous potential applications in areas ranging from gene therapy to population control. Some proposed applications involve CRISPR/Cas9 endonucleases integrated into an organisms genome, which raises questions about potentially harmful effects to the transgenic individuals. One application where this is particularly relevant are CRISPR-based gene drives, which promise a mechanism for rapid 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, designed to disentangle direct fitness costs due to the integration, expression, and target-site activity of Cas9 from costs due to potential off-target cleavage. Using a maximum likelihood framework, we find a moderate level of fitness costs due to off-target effects but do not detect significant direct costs. Costs of off-target effects are minimized 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. Our 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.

evolutionary biology↗

bioRxiv2020

Modeling CRISPR gene drives for suppression of invasive rodents

Invasive rodent populations pose a threat to biodiversity across the globe. When confronted with these new invaders, native species that evolved independently are often defenseless. CRISPR gene drive systems could provide a solution to this problem by spreading transgenes among invaders that induce population collapse. Such systems might be deployed even where traditional control methods are impractical or prohibitively expensive. Here, we develop a high-fidelity model of an island population of invasive rodents that includes three types of suppression gene drive systems. The individual-based model is spatially explicit and allows for overlapping generations and a fluctuating population size. Our model includes variables for drive fitness, efficiency, resistance allele formation rate, as well as a variety of ecological parameters. The computational burden of evaluating a model with such a high number of parameters presents a substantial barrier to a comprehensive understanding of its outcome space. We therefore accompany our population model with a meta-model that utilizes supervised machine learning to approximate the outcome space of the underlying model with a high degree of accuracy. This enables us to conduct an exhaustive inquiry of the population model, including variance-based sensitivity analyses using tens of millions of evaluations. Our results suggest that sufficiently capable gene drive systems have the potential to eliminate island populations of rodents under a wide range of demographic assumptions, but only if resistance can be kept to a minimal level. This study highlights the power of supervised machine learning for identifying the key parameters and processes that determine the population dynamics of a complex evolutionary system.

ecology↗

bioRxiv2019

Design and analysis of CRISPR-based underdominance toxin-antidote gene drives

CRISPR gene drive systems offer a mechanism for transmitting a desirable transgene throughout a population for purposes ranging from vector-borne disease control to invasive species suppression. In this simulation study, we assess the performance of several CRISPR-based underdominance gene drive constructs employing toxin-antidote principles. These drives disrupt the wild-type version of an essential gene using a CRISPR nuclease (the toxin) while simultaneously carrying a recoded version of the gene (the antidote). Drives of this nature allow for releases that could be potentially confined to a desired geographic location. This is because such drives have a nonzero invasion threshold frequency, referring to the critical frequency required for the drive to spread through the population. We model drives which target essential genes that are either haplosufficient or haplolethal, using nuclease promoters with expression restricted to the germline, promoters that additionally result in cleavage activity in the early embryo from maternal deposition, and promoters that have ubiquitous somatic expression. We also study several possible drive architectures, considering both "same-site" and "distant-site" systems, as well as several reciprocally targeting drives. Together, these drive variants provide a wide range of invasion threshold frequencies and options for both population modification and suppression. Our results suggest that CRISPR toxin-antidote underdominance drive systems could allow for the design of highly flexible and potentially confinable gene drive strategies.

genetics↗

bioRxiv2019

Suppression gene drive in continuous space can result in unstable persistence of both drive and wild-type alleles

Rapid evolutionary processes can produce drastically different outcomes when studied in panmictic population models versus spatial models where the rate of evolution is limited by dispersal. One such process is gene drive, which allows \"selfish\" genetic elements to quickly spread through a population. Engineered gene drive systems are being considered as a means for suppressing disease vector populations or invasive species. While laboratory experiments and modeling in panmictic populations have shown that such drives can rapidly eliminate a population, it is not yet clear how well these results translate to natural environments where individuals inhabit a continuous landscape. Using spatially explicit simulations, we show that instead of population elimination, release of a suppression drive can result in what we term \"chasing\" dynamics. This describes a condition in which wild-type individuals quickly recolonize areas where the drive has locally eliminated the population. Despite the drive subsequently chasing the wild-type allele into these newly re-colonized areas, complete population suppression often fails or is substantially delayed. This delay increases the likelihood that the drive becomes lost or that resistance evolves. We systematically analyze how chasing dynamics are influenced by the type of drive, its efficiency, fitness costs, as well as ecological and demographic factors such as the maximal growth rate of the population, the migration rate, and the level of inbreeding. We find that chasing is generally more common for lower efficiency drives and in populations with low dispersal. However, we further find that some drive mechanisms are substantially more prone to chasing behavior than others. Our results demonstrate that the population dynamics of suppression gene drives are determined by a complex interplay of genetic and ecological factors, highlighting the need for realistic spatial modeling to predict the outcome of drive releases in natural populations.

ecology↗

bioRxiv2019

Computational and experimental performance of CRISPR homing gene drive strategies with multiplexed gRNAs

CRISPR homing gene drives potentially have the capacity for large-scale population modification or suppression. However, resistance alleles formed by the drives can prevent them from successfully spreading. Such alleles have been found to form at high rates in most studies, including those in both insects and mammals. One possible solution to this issue is the use of multiple guide RNAs (gRNAs), thus allowing cleavage by the drive even if resistance sequences are present at some of the gRNA target sequences. Here, we develop a high-fidelity model incorporating several factors affecting the performance of drives with multiple gRNAs, including timing of cleavage, reduction in homology-directed repair efficiency due to imperfect homology around the cleavage site, Cas9 activity saturation, variance in the activity level of individual gRNAs, and formation of resistance alleles due to incomplete homology-directed repair. We parameterize the model using data from homing drive experiments designed to investigate these factors and then use it to analyze several types of homing gene drives. We find that each type of drive has an optimal number of gRNAs, usually between two and eight, dependent on drive type and performance parameters. Our model indicates that utilization of multiple gRNAs is insufficient for construction of successful gene drives, but that it provides a critical boost to drive efficiency when combined with other strategies for population modification or suppression.

genetics↗

bioRxiv2019

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

Engineered 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.

genetics↗

bioRxiv2019

A toxin-antidote CRISPR gene drive system for regional population modification

Engineered gene drives have been suggested as a mechanism for rapidly spreading genetic alterations through a population. One promising type of drive is the CRISPR homing drive, which has recently been demonstrated in several organisms. However, such drives face a major obstacle in the form of resistance against the drive that typically evolves rapidly. In addition, homing-type drives are generally self-sustaining, meaning that a drive would likely spread to all individuals of a species even when introduced at low frequency in a single location. Here, we develop a new form of CRISPR gene drive, the Toxin-Antidote Recessive Embryo (TARE) drive, which successfully limits resistance by targeting a recessive lethal gene while providing a recoded sequence to rescue only drive-carrying individuals. Our computational modeling shows that such a drive will have threshold-dependent dynamics, spreading only when introduced above a frequency threshold that depends on the fitness cost of the drive. We demonstrate such a drive in Drosophila with 88-95% transmission to the progeny of female drive heterozygotes. This drive was able to spread through a large cage population in just six generations following introduction at 24% frequency without any apparent evolution of resistance. Our results suggest that TARE drives constitute promising candidates for the development of effective, regionally confined population modification drives.

genetics↗