Mitigating Resistance to Gene Drive

Mabel Taracena, Ph.D. is a postdoctoral researcher at the Centers for Disease Control in Atlanta, Georgia where she is investigating RNAi based technologies for vector control. MORE ABOUT THE AUTHOR

A recent manuscript posted on bioRiv by Hammond et al (2018) report exciting improvements to CRISPR-based suppression gene drives in Anopheles gambiae, with the development of new CRISPR designs that reduce selection of resistance and results in much higher suppression outputs.

In the context of malaria transmission, vector control remains the frontline for disease elimination and gene drives are an especially appealing strategy due to the low costs, minimal infrastructure requirements, and the highly efficient results. However, as the force of selection for resistance to the drive has been proportional to the fitness cost imposed by it, previous attempts had resulted in the generation of mutations that eventually eliminated the gene drives.

Previous work of this group and others, allowed the identification of several key points to consider when designing gene drives, such as the functional constraint at the target site and the contribution of the end-joining repair pathways that can introduce mutations at the target site during the repair.

Image result for gene drive resistance

The basic mechanism of a ‘homing’ gene drive. If the endonuclease cannot cut at the homologous target site then that chromosome will be ‘resistant’ to invasion by the gene drive. There are a number of ways the homologous target site can become resistant to the gene drive. Minimizing the evolution of resistant target site alleles is important for achieving sustainable invasion of a gene drive into a population. Image credit:

In an elegant way to develop an improved system, they deliberately used a previously tested female fertility locus, and then exchanged the vasa promoter for novel germline promoters (zero population growth (zpg), nanos (nos) and exuperantia (exu)).  Promoters from these genes were selected because of their expression is restricted to the  germline.

This approach allowed a reliable quantification of any reduction in the occurrence or selection of resistant mutations. As a result, two very efficient new gene drives were obtained, as these not only restricted the spatio-temporal nuclease expression and but also dramatically improved the fitness and mitigated the development of resistance.

Furthermore, besides providing evidence that simple changes to the promoter controlling endonuclease expression can lead to drastic improvements in the rate of invasion of the gene drive, the experiments performed by Hammond et al. demonstrated that the initial mosquito release frequency has little impact upon the potential of the gene drive to spread within the population.

Anopheles gambiae

In as little as four generations the gene drive had spread to more than 97% of the population. To achieve these results, they developed a novel single generation resistance assay, designed to deplete non-functional mutations and reveal functional resistant mutations. This technique allowed a much more efficient separation of viable progeny with the right gene drives, which further potentiated the efficiency of their approach.

Overall, the contribution of this work to the CRISPR-based suppression gene drives is substantial, and will bring the strategy much closer to successful implementation.


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