THE MYXOMA VIRUS M-T4 GENE ENCODES A NOVEL RDEL-CONTAINING PROTEIN THAT IS RETAINED WITHIN THE ENDOPLASMIC-RETICULUM AND IS IMPORTANT FOR THE PRODUCTIVE INFECTION OF LYMPHOCYTES

To investigate the contribution of the myxoma virus M-T4 gene to viral virulence, both copies of the M-T4 gene were inactivated by disruptio n and insertion oi the Escherichia coil guanosine phosphoribosyltransf erase gene. Infection of European rabbits with the recombinant M-T4-de leted virus, vMyxlacT4(-), resulted in disease attenuation. In contras t, infection of rabbits with vMyxlac elicited the classical features o f lethal myxomatosis. A notable decrease in the number of secondary le sions in animals infected with vMyxlacT4(-) suggested an inability of the virus to disseminate in vivo. Infection of either a rabbit CD4(+) T cell line, RL-5, or primary rabbit peripheral blood lymphocytes with vMyxlacT4(-) resulted in the rapid induction of apoptosis. Sequence a nalysis of M-T4 revealed both an N-terminal signal sequence and a C-te rminal -RDEL sequence, suggesting that M-T4 resides in the endoplasmic reticulum. The M-T4 protein was found to be sensitive to endo H diges tion and confocal fluorescence microscopy demonstrated that M-T4 coloc alized with calreticulin, indicating that M-T4 is retained within the endoplasmic reticulum. Our results indicate that M-T4 is the first exa mple of an intracellular virulence factor in myxoma virus that functio ns from within the endoplasmic reticulum and is necessary for the prod uctive infection of lymphocytes. (C) 1997 Academic Press.