Marek's Disease Virus, a lymphotrophic, oncogenic alpha-herpesvirus of chickens, has historically presented a significant vaccine challenge to the poultry industry, and is also a useful model for herpesviral vaccine development. Herpesviruses impact both human and animal health, and the development of improved vaccines is needed to overcome obstacles such as a general lack of sterilizing immunity and the persistence of latent infection (both wild-type and vaccine viruses) in vaccinated hosts. Immune evasion by herpesviruses complicates vaccine development, as vaccine strains may evade multiple arms of immune recognition. Evasion of the major histocompatibility complex (MHC) class I has been recognized among a number of herpesviruses, including Marek's Disease Virus (MDV), and may interfere with CD8+ T cell recognition control of viral infection. Using plasmid-based gene cloning and in vitro transfection methods, the gene MDV012 was identified as an evasin of MHC class I, most likely inhibiting the TAP transporter. Potential knock-out viruses were constructed using BAC recombineering to introduce an A-to-C point mutation in the MDV012 start codon. These viruses grew more slowly and to lower titers than the wild-type virus. The MDV012ctg mutant gene was cloned from these viruses and tested in transfection assays. Surprisingly, the MDV012ctg mutant showed no difference in ability to down-regulate overall MHC class I expression relative to the wild-type MDV012, indicating that this gene is still translated or partially translated despite the mutation. Full knock-outs of MDV012 function will be required to test potentially non-MHC class I-evasive viruses.
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