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Drive is an expansion of a story of the same name that Sallis originally wrote for the noir anthology Measures of Poison (2002), published by Dennis McMillan Publications.[2] The novel was published by Poisoned Pen Press on September 1, 2005.[3]

Entertainment Weekly wrote that the novel "reads the way a Tarantino or Soderbergh neo-noir plays, artfully weaving through Driver's haunted memory and fueled by confident storytelling and keen observations about moviemaking, low-life living, and, yes, driving."[6]

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CRISPR gene drive systems allow the rapid spread of a genetic construct throughout a population. Such systems promise novel strategies for the management of vector-borne diseases and invasive species by suppressing a target population or modifying it with a desired trait. However, current homing-type drives have two potential shortcomings. First, they can be thwarted by the rapid evolution of resistance. Second, they lack any mechanism for confinement to a specific target population. In this study, we conduct a comprehensive performance assessment of several new types of CRISPR-based gene drive systems employing toxin-antidote (TA) principles, which should be less prone to resistance and allow for the confinement of drives to a target population due to invasion frequency thresholds.

The underlying principle of the proposed CRISPR toxin-antidote gene drives is to disrupt an essential target gene while also providing rescue by a recoded version of the target as part of the drive allele. Thus, drive alleles tend to remain viable, while wild-type targets are disrupted and often rendered nonviable, thereby increasing the relative frequency of the drive allele. Using individual-based simulations, we show that Toxin-Antidote Recessive Embryo (TARE) drives targeting an haplosufficient but essential gene (lethal when both copies are disrupted) can enable the design of robust, regionally confined population modification strategies with high flexibility in choosing promoters and targets. Toxin-Antidote Dominant Embryo (TADE) drives require a haplolethal target gene and a germline-restricted promoter, but they could permit faster regional population modification and even regionally confined population suppression. Toxin-Antidote Dominant Sperm (TADS) drives can be used for population modification or suppression. These drives are expected to spread rapidly and could employ a variety of promoters, but unlike TARE and TADE, they would not be regionally confined and also require highly specific target genes.

While very promising for suppression drives, such a strategy may be difficult to apply to population modification drives. This is because it relies on the principle that resistance alleles render the target gene nonfunctional, thereby enabling them to contribute to the overall goal of population suppression even if they slow the spread of the drive allele. A population modification drive, by contrast, would only be able to remain viable while removing resistance alleles if the drive targets an essential gene and itself contains a recoded version of the target gene that restores its function. This would require targeting a site that can be sufficiently recoded without rendering the target gene nonfunctional (i.e., the target sequence is not fully constrained). Yet at such a site, it should then also be possible for resistance alleles to maintain gene function [27].

Another inherent feature of homing drives that could limit their utility is the propensity to spread to distant populations with even small levels of migration [29], making it difficult to confine such a drive to a specific geographic region. This could be particularly undesirable for applications where the goal is suppression of invasive species or agricultural pests outside their native range [30]. Thus, new gene drive options are needed that are effective and flexible and can be confined to a target region.

A variety of TA systems are conceivable, depending on the nature of the target gene and the intended application of the drive (population modification or suppression). In this study, we provide a detailed discussion of several such systems, including those based on haplolethal genes and genes that are essential but haplosufficient (where disrupted alleles are recessive lethal), as well as specific genes required for sperm development.

The underlying design principle of all TA drive systems is that the drive alleles contain a toxin together with an antidote that rescues the effect of the toxin. We assume that the toxin is a CRISPR nuclease targeting an essential gene that will be disrupted and rendered nonfunctional when mutations are introduced at the cut sites through end-joining or homology-directed repair. The antidote consists of a recoded version of the gene, which does not match the gRNAs and therefore cannot be cleaved by the drive. Cells or individuals exposed to the toxin will often be nonviable, unless rescued by a drive allele. In contrast to homing drives that spread by directly increasing the number of drive alleles, TA drives spread by reducing the number of wild-type alleles (and thus still increasing the relative frequency of the drive). Various potential arrangements and targets for TA systems can be conceived. In this study, we will focus on three general classes of such systems:

TARE (Toxin-Antidote Recessive Embryo). These drives target an essential but haplosufficient gene. Disrupted alleles are recessive lethal (i.e., one functional copy of the gene is required for viability, which can be a drive or wild-type allele).

TADS (Toxin-Antidote Dominant Sperm). These drives target a gene that is transcribed in gametocytes after meiosis I in males, with this expression being critical for successful spermatogenesis. We assume that all sperm with a disrupted target allele are nonviable.

Figure 1a shows which genotypes are rendered nonviable in each of these classes of drive. The detailed features of these systems will be discussed in the relevant sections below. Generally, TARE systems are aimed for population modification while TADE and TADS systems can be used for both population modification and suppression. For TADE or TADS suppression, we assume that the drive and target loci are unlinked and that the drive is placed in an essential but haplosufficient fertility gene (that only affects fertility in one sex for TADE and specifically males for TADS), disrupting the gene with its presence (not by targeting it with gRNAs) so that drive homozygotes of one sex are sterile. We further discuss a population modification variant of TADE that we term Toxin-Antidote Double Dominant Embryo (TADDE) drive, which still targets a haplolethal gene but has a stronger rescue element such that a single drive allele is sufficient for an individual to be viable (Fig. 1a), allowing the drive to spread faster with a lower invasion threshold than TADE drive. Finally, we will discuss a variant of TADS where the drive is located on the Y chromosome (termed TADS Y-suppression). Such a system is expected to exhibit similar dynamics to previously studied X-shredder drives [41,42,43].

All of the TA systems we tested are able to spread quickly through the population in this idealized model (Figure S1), but most have frequency thresholds below which the drive will not invade if it carries a fitness cost (Fig. 1b). Realistic drives should carry at least a small fitness costs from drive components itself (from expression of the CRISPR nuclease, for example), and payload genes would likely add additional fitness costs (although these might be removed if mutations render the payload nonfunctional). When introduced above the threshold frequency, the drive is expected to increase in frequency and spread successfully, while below the frequency, it would likely be eliminated from the population. In contrast, TADS drives, as well as homing drives [4, 44] and X-shredders [44], all have a zero-threshold introduction frequency unless fitness costs are very high (drive homozygote fitness

TARE, TADDE, and Medea drives are not expected to go to fixation but instead reach equilibrium frequencies that are dependent on fitness costs (Fig. 1b). At equilibrium, all individuals are expected to carry at least one copy of the drive (Figure S1B), but some will carry disrupted alleles as well. Suppression forms of the drives are potentially capable of inducing high genetic loads (defined as the average net fitness reduction relative to a wild-type population of the same size after the drive reaches an equilibrium), though fitness costs can allow modification-type drives to induce a modest genetic load as well (Fig. 1d). However, loads based on such fitness costs in modification drives will usually be insufficient to eradicate a population or even substantially reduce its numbers, depending on ecological characteristics.

We will next study the individual TA systems more closely, exploring how their dynamics change as drive parameters are varied from the idealized model. The following analyses no longer assume a deterministic model of an infinite population as used in Fig. 1. Instead, they are based on our individual-based simulations, which seek to model a more realistic population of finite size with density regulation. These simulations therefore take stochastic effects into account, which can become particularly relevant for suppression approaches as population size decreases.

These drives constitute modification drives that target a gene that is essential and haplosufficient (disrupted alleles are recessive lethal), with the drive providing rescue (Fig. 2a). One consequence of this mechanism is that TARE drive will have threshold-dependent invasion dynamics (Figs. 1b and 2b). Another consequence is that embryo Cas9 cleavage from maternally deposited Cas9, which poses a major problem for homing-type drives, actually makes a TARE drive more efficient (Fig. 2c). For example, when a heterozygous female mates with a wild-type male, most of their offspring will end up carrying the drive. This is because those that did not inherit a drive allele from their mother will likely inherit a disrupted target allele, and the wild-type allele inherited from the father will then become disrupted due to maternal Cas9 activity, rendering those individuals nonviable. TARE drives should therefore be highly tolerant of variation in expression from the nuclease promoter. Indeed, the promoter of a TARE drive need not even be restricted to expression in the germline and early embryo. Constitutively active promoters would presumably work equally well (though they may have a higher fitness cost), as long as there is expression in germline or germline precursor cells. ff782bc1db