Mutant mice lacking p56lck, the T cell-specific protein tyrosine kinase, have a profound thymic atrophy and possess only an immature thymocyte population. Only 5 to 10% of normal levels of mature T cells (TCR-alpha beta+, CD4+, or CD8+) are present in these mice. These T cells, but also B cells of mutant mice, can be activated by mitogens, and T cells proliferate upon stimulation with IL-2 or mAb against the TCR. Thus, it appears that selected T cells can mature without lck and that they may have circumvented the need for this enzyme. This study evaluated the capacity of these mice to generate antiviral immune responses: 1) mutant mice failed to generate virus-specific CTL after acute infection with lymphocytic choriomeningitis virus (LCMV) or vaccinia virus. 2) LCMV was not cleared from the spleen on day 10 after infection. 3) Infection of LCMV into footpads of mutant mice did not induce a T cell-dependent swelling reaction. 4) Intracerebral inoculation with LCMV did not induce in mutant mice any clinical signs of sickness. 5) Vesicular stomatitis virus (VSV)-primed mutant mice could not eliminate nucleoprotein of VSV recombinant vaccinia virus upon intracerebral infection. 6) VSV infection of mutant mice elicited normal, specific T cell-independent IgM responses, but no significant switch to IgG. These results show that, despite the presence of in vitro activatable CD8+ and CD4+ T cells in the mutant mice, no biologically relevant functions can be detected. Therefore, these data indicate that p56lck is directly or indirectly crucially involved in in vivo Ag-induced T cell responses.