Inherently confinable split-drive systems in Drosophila

CRISPR-based gene-drive systems, which copy themselves via gene conversion mediated by the homology-directed repair (HDR) pathway, have the potential to revolutionize vector control. However, mutant alleles generated by the competing non-homologous end-joining (NHEJ) pathway, resistant to Cas9 cleavage, can interrupt the spread of gene-drive elements. We hypothesized that drives targeting genes essential for viability or reproduction also carrying recoded sequences that restore endogenous gene functionality should benefit from dominantly-acting maternal clearance of NHEJ alleles combined with recessive Mendelian culling processes. Here, we test split gene-drive (sGD) systems in Drosophila melanogaster that are inserted into essential genes required for viability (rab5, rab11, prosalpha2) or fertility (spo11). In single generation crosses, sGDs copy with variable efficiencies and display sex-biased transmission. In multigenerational cage trials, sGDs follow distinct drive trajectories reflecting their differential tendencies to induce target chromosome damage and/or lethal/sterile mosaic Cas9-dependent phenotypes, leading to inherently confinable drive outcomes. NHEJ alleles and Cas9 remnants after a gene drive introduction are scientific and public concerns. Here, the authors use split drives with recoded rescue elements to target essential genes and minimize the appearance of NHEJ alleles while also leaving no trace of Cas9.

Automated summary

CRISPR-based gene-drive systems have the potential to revolutionize vector control, but mutant alleles resistant to Cas9 cleavage can interrupt gene spread. Targeting essential genes with recoded rescue elements may reduce the appearance of NHEJ alleles and leave no trace of Cas9.