Scholarly Literature
This is a database of scholarly literature that concentrates currently on natural and engineered selfish genetic elements (gene drives). The latest are shown here.
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Spatial structure undermines parasite suppression by gene drive cargo
3896Bull, JJR, Christopher H.; Gomulkiewicz, Richard; Krone, Stephen M., PeerJ, 7:e7921. 2019-01-14 00:00:00.
Gene drives may be used in two ways to curtail vectored diseases. Both involve engineering the drive to spread in the vector population. One approach uses the drive to directly depress vector numbers, possibly to extinction. The other approach leaves intact the vector population but suppresses the disease agent during its interaction with the vector. This second application may use a drive engineered to carry a genetic cargo that blocks the disease agent. An advantage of the second application is that it is far less likely to select vector resistance to block the drive, but the disease agent may instead evolve resistance to the inhibitory cargo. However, some gene drives are expected to spread so fast and attain such high coverage in the vector population that, if the disease agent can evolve resistance only gradually, disease eradication may be feasible. Here we use simple models to show that spatial structure in the vector population can greatly facilitate persistence and evolution of resistance by the disease agent. We suggest simple approaches to avoid some types of spatial structure, but others may be intrinsic to the populations being challenged and difficult to overcome.
CRISPR in sub-Saharan Africa: Applications and education
3935Ogaugwu, CEA, S. O.; Adekoya, M. A., Trends in Biotechnology, 37:234-237. 2019-01-13 00:00:00.
Clustered regularly interspaced shortpalindromicrepeats (CRISPR) technology has enabled genetic engineering feats previously considered impracticable, offering great hopes for solutions to problems facing society. We consider it timely to highlight how CRISPR can benefit public health, medicine, and agriculture in sub-Saharan Africa (SSA) and offer recommendations for successful implementation.
Efficient allelic-drive in Drosophila
3915Guichard, AH, T.; Bobik, M.; Xu, X. R. S.; Klanseck, C.; Kushwah, R. B. S.; Berni, M.; Kaduskar, B.; Gantz, V. M.; Bier, E., Nature Communications, 10:1640. 2019-01-13 00:00:00.
Gene-drive systems developed in several organisms result in super-Mendelian inheritance of transgenic insertions. Here, we generalize this "active genetic" approach to preferentially transmit allelic variants (allelic-drive) resulting from only a single or a few nucleotide alterations. We test two configurations for allelic-drive: one, copy-cutting, in which a nonpreferred allele is selectively targeted for Cas9/guide RNA (gRNA) cleavage, and a more general approach, copy-grafting, that permits selective inheritance of a desired allele located in close proximity to the gRNA cut site. We also characterize a phenomenon we refer to as lethal-mosaicism that dominantly eliminates NHEJ-induced mutations and favors inheritance of functional cleavage-resistant alleles. These two efficient allelic-drive methods, enhanced by lethal mosaicism and a trans-generational drive process we refer to as "shadow-drive", have broad practical applications in improving health and agriculture and greatly extend the active genetics toolbox.
Engineered resistance to Zika virus in transgenic Aedes aegypti expressing a polycistronic cluster of synthetic small RNAs
3895Buchman, AG, S.; Li, M.; Antoshechkin, I.; Li, H. H.; Wang, H. W.; Chen, C. H.; Klein, M. J.; Duchemin, J. B.; Paradkar, P. N.; Akbari, O. S., Proceedings of the National Academy of Sciences of the United States of America, 116:3656-3661. 2019-01-13 00:00:00.
Recent Zika virus (ZIKV) outbreaks have highlighted the necessity for development of novel vector control strategies to combat arboviral transmission, including genetic versions of the sterile insect technique, artificial infection with Wolbachia to reduce population size and/or vectoring competency, and gene drive-based methods. Here, we describe the development of mosquitoes synthetically engineered to impede vector competence to ZIKV. We demonstrate that a polycistronic cluster of engineered synthetic small RNAs targeting ZIKV is expressed and fully processed in Aedes aegypti, ensuring the formation of mature synthetic small RNAs in the midgut where ZIKV resides in the early stages of infection. Critically, we demonstrate that engineered Ae. aegypti mosquitoes harboring the anti-ZIKV transgene have significantly reduced viral infection, dissemination, and transmission rates of ZIKV. Taken together, these compelling results provide a promising path forward for development of effective genetic-based ZIKV control strategies, which could potentially be extended to curtail other arboviruses.
Cleave and Rescue, a novel selfish genetic element and general strategy for gene drive
3934Oberhofer, GI, T.; Hay, B. A., Proceedings of the National Academy of Sciences of the United States of America, 116:6250-6259. 2019-01-12 00:00:00.
There is great interest in being able to spread beneficial traits throughout wild populations in ways that are self-sustaining. Here, we describe a chromosomal selfish genetic element, CleaveR [Cleave and Rescue (ClvR)], able to achieve this goal. ClvR comprises two linked chromosomal components. One, germline-expressed Cas9 and guide RNAs (gRNAs)-the Cleaver-cleaves and thereby disrupts endogenous copies of a gene whose product is essential. The other, a recoded version of the essential gene resistant to cleavage and gene conversion with cleaved copies-the Rescue-provides essential gene function. ClvR enhances its transmission, and that of linked genes, by creating conditions in which progeny lacking ClvR die because they have no functional copies of the essential gene. In contrast, thosewho inherit ClvR survive, resulting in an increase in ClvR frequency. ClvR is predicted to spread to fixation under diverse conditions. To test these predictions, we generated a ClvR element in Drosophilamelanogaster. ClvRtko is located on chromosome 3 and uses Cas9 and four gRNAs to disrupt melanogaster technical knockout (tko), an X-linked essential gene. Rescue activity is provided by tko from Drosophila virilis. ClvRtko results in germline and maternal carryover-dependent inactivation of melanogaster tko (> 99% per generation); lethality caused by this loss is rescued by the virilis transgene; ClvRtko activities are robust to genetic diversity in strains from five continents; and uncleavable but functional melanogaster tko alleles were not observed. Finally, ClvRtko spreads to transgene fixation. The simplicity of ClvR suggests it may be useful for altering populations in diverse species.
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9-mediated mutagenesis of the multiple edematous wings gene induces muscle weakness and flightlessness in Bactrocera dorsalis (Diptera: Tephritidae)
3954Zheng, WL, Q.; Sun, H.; Ali, M. W.; Zhang, H., Insect Molecular Biology, 28:222-234. 2019-01-12 00:00:00.
The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system is a versatile, efficient and heritable gene editing tool that can be useful for genome engineering. Bactrocera dorsalis (Hendel) is a major pest of agriculture that causes great economic losses. We used the B. dorsalis multiple edematous wings (Bdmew) gene as the target gene to explore the effectiveness of CRISPR/Cas9 for B. dorsalis genome manipulation. We studied the physiological functions of the Bdmew gene, particularly those related to muscle development. Site-specific genome editing was feasible using direct microinjection of specific guide RNA and the Cas9-plasmid into B. dorsalis embryos. Mutation frequencies ranged from 12.1 to 30.2% in the injected generation. Mosaic G0, with the mew mutation, was heritable to the next generation. The G1 displayed a series of defective phenotypes including muscle weakness, flightlessness, failure to eclose, wing folds and unbalanced movement. These results demonstrated that CRISPR/Cas9 can act as a highly specific, efficient, heritable tool for genome manipulation in B. dorsalis and this has significance for gene function research and genetic control of pests. The Bdmew gene possesses key functions in muscle development of B. dorsalis. Bdmew mutations cause a series of serious defects by interfering with muscle development and may provide a means for controlling B. dorsalis via a gene-based method such as gene drive.
CRISPR in Parasitology: Not Exactly Cut and Dried!
3894Bryant, JMB, S.; Glover, L.; Hutchinson, S.; Rachidi, N., Trends in Parasitology, 35:409-422. 2019-01-12 00:00:00.
CRISPR/Cas9 technology has been developing rapidly in the field of parasitology, allowing for the dissection of molecular processes with unprecedented efficiency. Optimization and implementation of a new technology like CRISPR, especially in nonmodel organisms, requires communication and collaboration throughout the field. Recently, a 'CRISPR in Parasitology' symposium was held at the Institut Pasteur Paris, bringing together scientists studying Leishmania, Plasmodium, Trypanosoma, and Anopheles. Here we share technological advances and challenges in using CRISPR/Cas9 in the parasite and vector systems that were discussed. As CRISPR/Cas9 continues to be applied to diverse parasite systems, the community should now focus on improvement and standardization of the technique as well as expanding the CRISPR toolkit to include Cas9 alternatives/derivatives for more advanced applications like genome-wide functional screens.
Super-Mendelian inheritance mediated by CRISPR-Cas9 in the female mouse germline
3914Grunwald, HAG, V. M.; Poplawski, G.; Xu, X. R. S.; Bier, E.; Cooper, K. L., Nature, 566:105-109. 2019-01-12 00:00:00.
A gene drive biases the transmission of one of the two copies of a gene such that it is inherited more frequently than by random segregation. Highly efficient gene drive systems have recently been developed in insects, which leverage the sequence-targeted DNA cleavage activity of CRISPR-Cas9 and endogenous homology-directed repair mechanisms to convert heterozygous genotypes to homozygosity(1-4). If implemented in laboratory rodents, similar systems would enable the rapid assembly of currently impractical genotypes that involve multiple homozygous genes (for example, to model multigenic human diseases). To our knowledge, however, such a system has not yet been demonstrated in mammals. Here we use an active genetic element that encodes a guide RNA, which is embedded in the mouse tyrosinase (Tyr) gene, to evaluate whether targeted gene conversion can occur when CRISPR-Cas9 is active in the early embryo or in the developing germline. Although Cas9 efficiently induces double-stranded DNA breaks in the early embryo and male germline, these breaks are not corrected by homology-directed repair. By contrast, Cas9 expression limited to the female germline induces double-stranded breaks that are corrected by homology-directed repair, which copies the active genetic element from the donor to the receiver chromosome and increases its rate of inheritance in the next generation. These results demonstrate the feasibility of CRISPR-Cas9-mediated systems that bias inheritance of desired alleles in mice and that have the potential to transform the use of rodent models in basic and biomedical research.
CRISPR-Cas9. The greatest advancement in genetic edition techniques requires an ethical reflection
3913Gomez-Tatay, LA, J., Cuadernos De Bioetica, 30:171-185. 2019-01-11 00:00:00.
The adaptation of the CRISPR system as a genetic editing tool has led to a revolution in many fields of application, as this technique is considerably faster, easier to perform and more efficient than predecessor techniques. However, some of these applications raise objective ethical issues that must be addressed. In this paper we discuss, based on the most recent data, the different issues related to CRISPR applications on the germ line, its introduction in clinical trials, the genetic edition of animals and plants for human consumption and the novel gene drive.
Efficient somatic and germline genome engineering of Bactrocera dorsalis by the CRISPR/Cas9 system
3953Zhao, STX, Z. Z.; Liu, Z. G.; Liu, Y. H.; Liu, X. R.; Chen, Z.; Li, J. H.; Yan, R. H., Pest Management Science, 75:1921-1932. 2019-01-11 00:00:00.
Bactrocera dorsalis (Hendel), a very destructive insect pest of many fruits and vegetables, is widespread in many Asian countries. To facilitate control of this pest, it is essential to investigate its genetics and gene function using targeted gene disruption. RESULTS Here, we describe successful targeted mutagenesis of the white and transformer genes in B. dorsalis through use of the clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) system. Co-injection of the white sgRNA and Cas9 mRNA into B. dorsalis embryos caused eye color change, and the white mutations in the germline were heritable. CRISPR-mediated knockout of the sex determination gene transformer (tra) in B. dorsalis resulted in a male-biased sex ratio and adult flies with abnormal outer and interior reproductive organs. Small indels and substitutions were induced by CRIRPR for both genes. CONCLUSION Our data demonstrate that somatic and germline genome engineering of the pest B. dorsalis can be performed efficiently using the CRISPR/Cas9 system, opening the door to the use of the CRISPR-mediated method for functional annotations of genes in B. dorsalis and for its population control using, for example, such as gene drive. (c) 2018 Society of Chemical Industry
Modelling the potential of genetic control of malaria mosquitoes at national scale
3933North, ARB, Austin; Godfray, H. Charles J., BMC Biology, 17:26. 2019-01-11 00:00:00.
The persistence of malaria in large parts of sub-Saharan Africa has motivated the development of novel tools to complement existing control programmes, including gene-drive technologies to modify mosquito vector populations. Here, we use a stochastic simulation model to explore the potential of using a driving-Y chromosome to suppress vector populations in a 106 km2 area of West Africa including all of Burkina Faso.
The EU regulatory framework on genetically modified organisms (GMOs)
3893Bruetschy, C, Transgenic Research, 28:169-174. 2019-01-11 00:00:00.
The European Union (EU) legislation on genetically modified organisms (GMOs) aims to ensure a high level of protection for human, animal and environmental health and a well-functioning EU internal market. The framework regulates the release of GMOs into the environment and their use as, or in, food and feed. It has three main pillars: pre-market authorisation based on a prior risk assessment, traceability and labelling. Within this legal framework, the EU has authorised the placing on the market of 118 GMOs so far. These have been obtained through long-standing techniques of genetic modification, namely transgenesis. Following the adoption of the GMO legislation, new techniques of genetic modification, including new mutagenesis techniques, have been developed, which have raised questions regarding the applicability of the GMO legislation and attracted a lot of attention from stakeholders and the general public. This article provides an overview of EU GMO legislation and implementation of the EU Court of Justice ruling on organisms obtained by mutagenesis techniques, issued in July 2018. It also updates on the recent initiatives by the European Commission and EU Member States on new developments in biotechnology. The manuscript is based on the author's contribution at the OECD Conference on Genome Editing, Applications in Agriculture, Implications for Health, Environment and Regulation held in Paris on 28-29 June 2018. It is complemented with updated information.
A genetic system for biasing the sex ratio in mice
3952Yosef, IEB, Liat; Globus, Rea; Shlomovitz, Inbar; Munitz, Ariel; Gerlic, Motti; Qimron, Udi, EMBO reports, 20:e48269. 2019-01-10 00:00:00.
Biasing the sex ratio of populations of different organisms, including plants, insects, crustacean, and fish, has been demonstrated by genetic and non-genetic approaches. However, biasing the sex ratio of mammalian populations has not been demonstrated genetically. Here, we provide a first proof of concept for such a genetic system in mammals by crossing two genetically engineered mouse lines. The maternal line encodes a functional Cas9 protein on an autosomal chromosome, whereas the paternal line encodes guide RNAs on the Y chromosome targeting vital mouse genes. After fertilization, the presence of both the Y-encoded guide RNAs from the paternal sperm and the Cas9 protein from the maternal egg targets the vital genes in males. We show that these genes are specifically targeted in males and that this breeding consequently self-destructs solely males. Our results pave the way for a genetic system that allows biased sex production of livestock.
Daisy-chain gene drives for the alteration of local populations
3932Noble, CM, J.; Olejarz, J.; Buchthal, J.; Chavez, A.; Smidler, A. L.; DeBenedictis, E. A.; Church, G. M.; Nowak, M. A.; Esvelt, K. M., Proceedings of the National Academy of Sciences of the United States of America, 116:8275-8282. 2019-01-10 00:00:00.
If they are able to spread in wild populations, CRISPR-based gene-drive elements would provide new ways to address ecological problems by altering the traits of wild organisms, but the potential for uncontrolled spread tremendously complicates ethical development and use. Here, we detail a self-exhausting form of CRISPR-based drive system comprising genetic elements arranged in a daisy chain such that each drives the next. "Daisydrive" systems can locally duplicate any effect achievable by using an equivalent self-propagating drive system, but their capacity to spread is limited by the successive loss of nondriving elements from one end of the chain. Releasing daisy-drive organisms constituting a small fraction of the local wild population can drive a useful genetic element nearly to local fixation for a wide range of fitness parameters without self-propagating spread. We additionally report numerous highly active guide RNA sequences sharing minimal homology that may enable evolutionarily stable daisy drive as well as self-propagating CRISPR-based gene drive. Especially when combined with threshold dependence, daisy drives could simplify decision-making and promote ethical use by enabling local communities to decide whether, when, and how to alter local ecosystems.
Precision control of CRISPR-Cas9 using small molecules and light
3912Gangopadhyay, SAC, K. J.; Manna, D.; Lim, D.; Maji, B.; Zhou, Q. X.; Choudhary, A., Biochemistry, 58:234-244. 2019-01-10 00:00:00.
The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas system is an adaptive immune system of bacteria that has furnished several RNA-guided DNA endonucleases (e.g., Cas9) that are revolutionizing the field of genome engineering. Cas9 is being used to effect genomic alterations as well as in gene drives, where a particular trait may be propagated through a targeted species population over several generations. The ease of targeting catalytically impaired Cas9 to any genomic loci has led to development of technologies for base editing, chromatin imaging and modeling, epigenetic editing, and gene regulation. Unsurprisingly, Cas9 is being developed for numerous applications in biotechnology and biomedical research and as a gene therapy agent for multiple pathologies. There is a need for precise control of Cas9 activity over several dimensions, including those of dose, time, and space in these applications. Such precision controls, which are required of therapeutic agents, are particularly important for Cas9 as off-target effects, chromosomal translocations, immunogenic response, genotoxicity, and embryonic mosaicism are observed at elevated levels and with prolonged activity of Cas9. Here, we provide a perspective on advances in the precision control of Cas9 over aforementioned dimensions using external stimuli (e.g., small molecules or light) for controlled activation, inhibition, or degradation of Cas9.
Promises and perils of gene drives: Navigating the communication of complex, post-normal science
3892Brossard, DB, Pam; Gould, Fred; Wirz, Christopher D., Proceedings of the National Academy of Sciences of the United States of America, 116:7692-7697. 2019-01-10 00:00:00.
In November of 2017, an interdisciplinary panel discussed the complexities of gene drive applications as part of the third Sackler Colloquium on “The Science of Science Communication.” The panel brought together a social scientist, life scientist, and journalist to discuss the issue from each of their unique perspectives. This paper builds on the ideas and conversations from the session to provide a more nuanced discussion about the context surrounding responsible communication and decision-making for cases of post-normal science. Deciding to use gene drives to control and suppress pests will involve more than a technical assessment of the risks involved, and responsible decision-making regarding their use will require concerted efforts from multiple actors. We provide a review of gene drives and their potential applications, as well as the role of journalists in communicating the extent of uncertainties around specific projects. We also discuss the roles of public opinion and online environments in public engagement with scientific processes. We conclude with specific recommendations about how to address current challenges and foster more effective communication and decisionmaking for complex, post-normal issues, such as gene drives.
CRISPR ethics: Moral considerations for applications of a powerful tool
3891Brokowski, C. and Adli, M., Journal of Molecular Biology, 431:88-101. 2019-01-09 00:00:00.
With the emergence of CRISPR technology, targeted editing of a wide variety of genomes is no longer an abstract hypothetical, but occurs regularly. As application areas of CRISPR are exceeding beyond research and biomedical therapies, new and existing ethical concerns abound throughout the global community about the appropriate scope of the systems' use. Here we review fundamental ethical issues including the following: 1) the extent to which CRISPR use should be permitted; 2) access to CRISPR applications; 3) whether a regulatory framework(s) for clinical research involving human subjects might accommodate all types of human genome editing, including editing of the germline; and 4) whether international regulations governing inappropriate CRISPR utilization should be crafted and publicized. We conclude that moral decision making should evolve as the science of genomic engineering advances and hold that it would be reasonable for national and supranational legislatures to consider evidence-based regulation of certain CRISPR applications for the betterment of human health and progress. (C) 2018 Published by Elsevier Ltd.
CRISPR-Clear: A fieldable detection procedure for potential CRISPR-Cas9 gene drive based bioweapons.
3931Nieuwenweg, ACvG, Martijn M.; Horsting, Angelina; Hegge, Jorrit W; Velders, Aldrik; Saggiomo, Vittorio, ChemRxiv, 2019:1-11. 2019-01-09 00:00:00.
Rapid progression in genetic modification research has made gene editing increasingly cheaper and easier to use. CRISPR-Cas9 for example, allows for the specific alteration of the genome of an organism with relative simplicity and low costs. This raised a worrying question; can genetic modification techniques be used to create novel bioweapons? A specific scenario is the initiation of a synthetic gene drive for malicious purposes. A synthetic gene drive can be used to quickly spread a mutation through an entire population. This mutation could alter vectors in such a way that they will spread human diseases or eradicate essential organisms. Since a gene drive spreads efficiently through a population, timely detection is essential. Thus, a quick and field deployable screening method is needed to counteract the malicious use of gene drives.; ; Here, we show a battery-operated, sensitive screening method, named CRISPR-Clear, for the detection of gene drive modified organisms. CRISPR-Clear is based on the combination of three components: 1) A DNA amplification technique known as loop-mediated isothermal amplification (LAMP) for detecting the presence of a gene drive; b) a portable battery-operated Arduino device which heats up the sample to allow DNA amplification, and c) a naked-eye visualization of the results.; ; We designed and tested six LAMP primers targeting a Cas9 endonuclease-based gene drive, assembled a battery-operated Arduino device and tested the naked-eye visualization method. In addition, we were able to detect the presence of the Cas9 gene, extracted from a transformed bacteria, providing a proof-of-concept of the CRISPR-Clear device.
Gene drives as a new quality in GMO releases-a comparative technology characterization
3911Friess, JLvG, A.; Giese, B., Peerj, 7:e6793. 2019-01-09 00:00:00.
Compared to previous releases of genetically modified organisms (GMOs) which were primarily plants, gene drives represent a paradigm shift in the handling of GMOs: Current regulation of the release of GMOs assumes that for specific periods of time a certain amount of GMOs will be released in a particular region. However, now a type of genetic technology arises whose innermost principle lies in exceeding these limits-the transformation or even eradication of wild populations. The invasive character of gene drives demands a thorough analysis of their functionalities, reliability and potential impact. But such investigations are hindered by the fact that an experimental field test would hardly be reversible. Therefore, an appropriate prospective assessment is of utmost importance for an estimation of the risk potential associated with the application of gene drives. This work is meant to support the inevitable characterization of gene drives by a comparative approach of prospective technology assessment with a focus on potential sources of risk. Therein, the hazard and exposure potential as well as uncertainties with regard to the performance of synthetic gene drives are addressed. Moreover, a quantitative analysis of their invasiveness should enable a differentiated evaluation of their power to transform wild populations.
Stakeholder attitudes towards the use of recombinant technology to manage the impact of an invasive species: Sea Lamprey in the North American Great Lakes
3951Thresher, REJ, M.; Drake, D. A. R., Biological Invasions, 21:575-586. 2019-01-09 00:00:00.
Several factors, including: (1) on-going difficulties of cost-effectively managing invasive species; (2) recent successes in using recombinant genetics to suppress mosquito populations; and, (3) developments in gene-drive technology, have re-invigorated interest in using genetic biotechnology to manage the impacts of invasive species. However, the extent to which there is social license' to develop and use these technologies has not been widely canvassed. We surveyed stakeholders involved directly and indirectly in managing Sea Lamprey (Petromyzon marinus) in the upper North American Great Lakes and a key community group of resource usersrecreational fishersto assess their support and concerns about researching, developing, and potentially implementing recombinant methods that an expert group assessed as likely to be effective in managing Sea Lamprey in the Great Lakes. Both groups overwhelmingly supported initiating R&D and, if risks were deemed very low, undertaking steps towards implementation. The key concern expressed by both groups was the risk of impacts to non-target taxa, including valued native populations of Sea Lamprey outside of the Great Lakes. Few respondents expressed opposition based on ethical or moral grounds, which contrasts with previous surveys on the use of recombinant technology in general. The broad support for R&D into recombinant approaches is likely to reflect trust in the nominated implementing agency (the Great Lakes Fishery Commission), its history of extensive consultation prior to undertaking management actions, and the hope that genetic biocontrol could solve the Sea Lamprey problem rather than simply managing it.
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