Role of the C-terminal RDEL motif of the myxoma virus M-T4 protein in terms of apoptosis regulation and viral pathogenesis

The purpose of this study was to investigate the significance of the termin al RDEL motif of the myxoma virus M-T4 protein in terms of apoptosis regula tion and rate in viral virulence. To accomplish this, a recombinant myxoma virus was created in which the C-terminal RDEL motif of M-T4 was deleted an d a selectable marker (Ecogpt) was inserted immediately downstream. We hypo thesized that removal of the RDEL motif from M-T4 would alter the subcellul ar localization of the protein and provide insight into its antiapoptotic r ole. Surprisingly, removal of the RDEL motif from M-T4 did not affect local ization of the protein within the endoplasmic reticulum (ER), but it did re duce the stability of the mutant protein. Pulse-chase immunoprecipitation a nd endoglycosidase H analysis coupled with confocal fluorescent light micro scopy demonstrated that the M-T4 RDEL- mutant protein is retained in the ER like wildtype M-T4 and suggests that the C-terminal RDEL motif is not the sole determinant for M-T4 localization to the ER. Infection of cultured rab bit lymphocytes with the M-T4 RDEL- mutant virus results in an intermediate apoptosis phenotype compared with the wildtype and M-T4 knockout mutant vi ruses, A novel myxomatosis phenotype was observed in European rabbits when infected with the recombinant M-T4 RDEL- mutant virus. Rabbits infected wit h the M-T4 RDEL- virus on day 9 postinfection exhibited an exacerbated edem atous and inflammatory response at secondary sites of infections, particula rly the ears. Our results indicate that the C-terminal RDEL motif may not h e solely responsible for retention of M-T4 to the ER and that M-T4 may have a dual function In protecting infected lymphocytes from apoptosis end in m odulating the inflammatory response to virus infection. (C) 1999 Academic P ress.