C3H mouse mammary tumor virus superantigen function requires a splice donor site in the envelope gene

Mouse mammary tumor virus (MMTV) encodes a superantigen (Sag) that is requi red for efficient milk-borne transmission of virus from mothers to offsprin g. The mRNA used for Sag expression is controversial, and at least four dif ferent promoters (two in the long terminal repeat and two in the envelope g ene) for sag mRNA have been reported. To determine which RNA is responsible for Sag function during milk-borne MMTV transmission, we mutated a splice donor site unique to a spliced sag RNA from the 5' envelope promoter. The s plice donor mutation in an infectious provirus was transfected into XC cell s and injected into BALB/c mice. Mice injected with wild-type provirus show ed Sag activity by the deletion of Sag-specific T cells and induction of ma mmary tumors in 100% of injected animals. However, mice injected with the s plice donor mutant gave sporadic and delayed T-cell deletion and a low perc entage of mammary tumors with a long Latency, suggesting that the resulting tumors were due to the generation of recombinants with endogenous MMTVs. T hird-litter offspring of mice injected with wild-type provirus showed Sag-s pecific T-cell deletion and developed mammary tumors with kinetics similar to those for mice infected by nursing on MMTV-infected mothers, whereas the third-litter offspring of the splice donor mutant-injected mice did not. O ne of the fifth-litter progeny of splice donor mutant-injected mice showed C3H Sag activity and had recombinants that repaired the splice donor mutati on, thus confirming the necessity for the splice donor site for Sag functio n. These experiments are the first to show that the spliced sag mRNA from t he 5' envelope promoter is required for efficient milk-borne transmission o f C3H MMTV.