Tumor-secreted gp96-Ig is highly immunogenic and triggers CD8 T cell-mediated tumor rejection. In vivo secreted gp96-Ig and gp96-myc cause NK activation and clonal expansion of specific CD8(+) CTL in wild-type and in Fas-ligand-deficient (gld) mice but not in perforin- (PKO) or IFN-gamma-deficient (GKO) mice. Transfer of perforin-competent NK cells restores the ability of PKO mice to clonally expand CD8 CTL in response to gp96-Ig. The data demonstrate an essential role for perforin-mediated functions in the activation of innate and adaptive immunity by heat shock protein gp96-peptide complexes. Crosspresentation of antigens by heat shock proteins seems to require a perforin-dependent positive feedback loop between NK and DC for both sustained NK activation and clonal CTL expansion. The studies also explain how depressed NK activity in patients with tumors or after viral infections could diminish CTL responses.