Abstract
Avian leukosis viruses (ALVs), which are pathogens of concern in domestic poultry, utilize specific receptor proteins for cell entry that are both necessary and sufficient for host susceptibility to a given ALV subgroup. This unequivocal relationship offers receptors as suitable targets of selection and biotechnological manipulation with the aim of obtaining virus-resistant poultry. This approach is further supported by the existence of natural knock-outs of receptor genes that segregate in inbred lines of chickens. We used CRISPR/Cas9 genome editing tools to introduce frame-shifting indel mutations into tva, tvc, and tvj loci encoding receptors for the A, C, and J ALV subgroups, respectively. For all three loci, the homozygous frame-shifting indels generating premature stop codons induced phenotypes which were fully resistant to the virus of respective subgroup. In the tvj locus, we also obtained in-frame deletions corroborating the importance of W38 and the four amino-acids preceding it. We demonstrate that CRISPR/Cas9-mediated knock-out or the fine editing of ALV receptor genes might be the first step in the development of virus-resistant chickens.
Highlights
Avian leukosis viruses (ALV) have been recognized as the cause of serious commercial losses in poultry industry since the 1920s
Results of the assay for each of the CRISPR/Cas9 constructs are shown as agarose electrophoresis controlled by markers of molecular size and wild-type DF-1 are shown cells.as agarose electrophoresis controlled by markers of molecular size and wild-type DF-1 cells
Our results clearly demonstrate the dependence of virus entry on intact sequences of respective receptors for the ALV subgroups A, C, and J
Summary
Avian leukosis viruses (ALV) have been recognized as the cause of serious commercial losses in poultry industry since the 1920s. The ALV subgroups have traditionally been classified by their range of susceptible/resistant hosts, antibody cross-neutralization, and interference in superinfection experiments [3]. The discovery of subgroup-specific receptors has explained the molecular mechanism of genetic susceptibility to ALVs [4]. A protein of the low-density lipoprotein receptor family [5,6], serves as the receptor for the ALV-A subgroup. Subgroups B, D, and E utilize the same Tvb receptor, a tumor necrosis factor receptor-related protein [7,8,9]. Subgroup C ASLVs enters cells via the Tvc protein of the butyrophilin family, which consists two immunoglobulin-like domains [10]. Tva, Tvb, Viruses 2018, 10, 605; doi:10.3390/v10110605 www.mdpi.com/journal/viruses
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