TALENs-mediated homozygous CCR5A32 mutations endow CD4+ U87 cells with resistance against HIV-1 infection

Abstract

Since evidence suggests that transplantation of bone marrow stem cells with the C-C chemokine receptor type 5 (CCR5)Δ32/Δ32 genotype may cure patients infected with human immunodeficiency virus (HIV)-1, the present study aimed to reproduce the CCR5Δ32 mutation in cluster of differentiation (CD)4+ U87 cells using genome engineering methods. A modified transcription activator-like effector nucleases (TALENs) technique, combined with homologous recombination for site-specific, size-controlled and homozygous DNA deletions, was used to reproduce the homozygous CCR5Δ32 mutation in CD4+ U87 cells. The results indicated that the frequency of the TALENs-targeted mutation reached 50.4% without any selection, whereas homologous recombination from CCR5 to CCR5Δ32 occurred in 8.8% of targeted cells. Notably, a HIV-1 challenge test demonstrated that CCR5Δ32/Δ32 CD4+ U87 cells were resistant to HIV infection. In conclusion, engineered CCR5Δ32/Δ32 mutations endowed CD4+ U87 cells with resistance against HIV-1 infection; this site-specific, size-controlled and homozygous DNA deletion technique was able to induce precise genomic editing, i.e., the deletion or insertion of a predetermined length of DNA sequence at a specific locus throughout the genome.