CD8(+) lymphocytes are critical to the control and elimination of viral pathogens. Impaired CD8(+) responses are well recognized in lentiviral infections; however, the mechanisms underlying CD8(+) impairment remain elusive. Using the feline immunodeficiency virus (FIV) model for human AIDS, we reported previously that CD4(+)CD25(+) Treg cells in both the acute and long-term, asymptomatic phase of infection are constitutively activated and suppress CD4(+)CD25(-) T cell responses. In the current study, we have demonstrated that CD4(+)CD25(+) Treg cells suppress CD8(+) responses to immune stimulation during both the acute and chronic, asymptomatic phase of FIV infection and that the mechanism of suppression may be mediated by membrane-associated TGF-beta (mTGF-beta) on CD4(+)CD25(+) lymphocytes. Depletion of CD4(+)CD25(+) lymphocytes from lymph node suspensions significantly enhanced production of IFN-gamma during the acute phase of infection and coculture of CD8(+) lymphocytes with CD4(+)CD25(+) lymphocytes resulted in suppression of CD8(+) IFN-gamma during both the acute and chronic stages of infection. FACS analysis indicated that there was TGF-betaRII upregulation on CD8(+) cells from FIV(+) cats during the acute and chronic stage of infection. In addition, there was upregulation of mTGF-beta on the CD4(+)CD25(+) subset in chronically infected cats. In support of activation of the TGF-beta signaling pathway, Western blotting showed Smad 2 phosphorylation in CD8(+) targets following CD4(+)CD25(+)/CD8(+) coculture. These results demonstrate the suppressive effect CD4(+)CD25(+) Treg cells have on the CD8(+) immune response during the acute and chronic stages of FIV infection and suggest that the mechanism of suppression may be mediated by mTGF-beta.