Immunization of C,H mice with a vaccinia virus (Vv) recombinant expressing the measles virus (MV) nucleoprotein (NP) induces an H-2K(k)-restricted cytotoxic T lymphocyte (CTL) response. With reference to the predicted peptide epitope motifs binding to this MHC class I molecule, we have used synthetic peptides derived from the primary sequence of the MV NP to establish the identity of the Kk-restricted CTL epitopes. Two octameric peptides, LDRLVRLI (aa 52-59) and VESPGQLI (aa 81-88) sensitized P815-K-k cells to lysis by MV NP-induced CTLs. In contrast to LDRLVRLI, the sequence VESPGQLI is also present in the primary sequence of the NP from the closely related canine distemper virus (CDV). In vitro stimulation of spleen cells from W NP-immunized mice with the peptides showed that peptide VESPGQLI induced CTLs which could also lyse CDV-infected cells, whereas peptide LDRLVRLI could only lyse cells presenting the MV protein. Different concentrations of peptides were used, and the lysis efficiencies for both epitopes were shown to be of the same order. The value of predicative motifs for determining MHC class I CTL epitopes is discussed, (C) 1994 Academic Press, Inc.