Isolation of a Marek's disease virus (MDV) glycoprotein C-negative mutant and regulation of major histocompatibility complex molecules during MDV infection

A Marek's disease virus (MDV) mutant was constructed that contains the lac Z and xanthine-guanine phosphoribosyltransferase (gpt) genes of E. coli inserted into the glycoprotein C (gC, A antigen, gp57–65) homolog gene. This mutant (RB1BgC gptlac) had the expected genomic structure and failed to produce a 1.8-kb gC-specific transcript in RB1B-infected chicken embryo fibroblasts (CEF). The tumor incidence in RB1BgCgptlac-inoculated chickens (2/16) was considerably lower than that seen in chickens inoculated with the RB1B parent (19/32). No chickens exposed to RB1BgCgptlac by contact (0/22) developed tumors; whereas, tumors were readily apparent in chickens exposed by contact to the parent virus (12/24). Attempts to isolate a gC rescuant both in culture and in vivo were unsuccessful possibly due to instability of the green fluorescent protein (GFP) or lowered transfection efficiency of the RB1BgCgptlac strain.; Several herpesviruses, including herpes simplex virus and cytomegalovirus, are able to evade immune recognition by down-regulating the cell-surface expression of MHC class I molecules. We compared steady-state RNA levels by microarrays and northern analyses and found slightly induced levels of MHC class I, MHC class II and β₂-microglobulin mRNAs in RB1B-infected CEF. Cell surface expression of the MHC was examined in the presence and absence of an RB1B infection at 8, 24, 48, 96, and 144 hours post-infection using flow cytometry. For at least one time point, levels of MHC class I and β ₂-microglobulin were higher in RB1B-infected CEF. Upon closer examination using confocal images, we showed that individual virus-infected cells expressed lower levels of MHC class I on the surface, but uninfected cells within the infected culture expressed high levels of surface MHC class I. Supernatants collected from RB1B-infected cultures 96 hours post-infection, when exposed to fresh uninfected cells, induced a significant increase in the number of cells expressing MHC class I and β₂-microglobulin on the cell surface. MDV infection appears to down-regulate MHC class I cell-surface expression in individual infected cells, but produces a soluble factor that can increase MHC class I and β₂-microglobulin expression in uninfected cells present in the culture.