Overcoming evolved resistance to population-suppressing homing-based gene drives.
Overcoming evolved resistance to population-suppressing homing-based gene drives.
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DOI：
10.1038/s41598-017-02744-7
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发表时间：
2017-06-19
影响因子：
4.6
通讯作者：
Akbari OS
中科院分区：
文献类型：
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作者：
Marshall JM;Buchman A;Sánchez C HM;Akbari OS
最近开发的基于 CRISPR-Cas9 的用于抑制冈比亚按蚊的归巢系统令人鼓舞；然而，根据目前的设计，归巢抗性等位基因的缓慢出现预计将导致受抑制的种群迅速反弹，因为归巢抗性等位基因比功能性、种群抑制性归巢等位基因具有显着的适应性优势。为了探讨这个问题，我们开发了一个数学模型来估计归巢抗性等位基因产生的耐受率，以抑制给定大小的野生种群。我们的结果表明，为了实现有意义的群体抑制，抗性等位基因产生的容忍率比基于 CRISPR-Cas9 的归巢系统的当前设计观察到的要小几个数量级。为了解决这个问题，我们从理论上探索了一种导引RNA（gRNA）多重的归巢系统架构，提高了有效归巢率并降低了有效抗性等位基因生成率。建模结果表明，可抑制的种群规模随着多重 gRNA 的数量呈指数增长，并且通过四种多重 gRNA，可能会在大陆范围内抑制蚊子物种。我们还展示了成功地使用多重核酶侧翼 gRNA 来诱导果蝇体内突变的原理验证——这种策略可以很容易地应用于在相关生物体中设计稳定的、基于归巢的驱动器。
The recent development of a CRISPR-Cas9-based homing system for the suppression of Anopheles gambiae is encouraging; however, with current designs, the slow emergence of homing-resistant alleles is expected to result in suppressed populations rapidly rebounding, as homing-resistant alleles have a significant fitness advantage over functional, population-suppressing homing alleles. To explore this concern, we develop a mathematical model to estimate tolerable rates of homing-resistant allele generation to suppress a wild population of a given size. Our results suggest that, to achieve meaningful population suppression, tolerable rates of resistance allele generation are orders of magnitude smaller than those observed for current designs for CRISPR-Cas9-based homing systems. To remedy this, we theoretically explore a homing system architecture in which guide RNAs (gRNAs) are multiplexed, increasing the effective homing rate and decreasing the effective resistant allele generation rate. Modeling results suggest that the size of the population that can be suppressed increases exponentially with the number of multiplexed gRNAs and that, with four multiplexed gRNAs, a mosquito species could potentially be suppressed on a continental scale. We also demonstrate successful proof-of-principle use of multiplexed ribozyme flanked gRNAs to induce mutations in vivo in Drosophila melanogaster – a strategy that could readily be adapted to engineer stable, homing-based drives in relevant organisms.