Infection with human T-cell leukemia virus type-I (HTLV-1) is associated with a low incidence of morbidity in the form of adult T-cell leukemia as well as neurologic disease, including tropical spastic paraparesis/HTLV-I-associated myelopathy, suggesting that there are other important factors which determine outcome of infection. HTLV-I and the human herpesvirus, cytomegalovirus (HCMV), have both been shown to infect OKT4+ T lymphocytes in vitro as well as in vivo. We investigated the effects of expression of HCMV IE-2 protein(s) on the HTLV-I long terminal repeat (LTR) containing the promoter elements in T-cell lines and primary lymphocytes. A consistent repressive effect was observed on HTLV-I LTR-driven chloramphenicol acetyl transferase activity after cotransfection with the HCMV IE-2 gene region, both in HTLV-I-producing cell lines as well as in uninfected primary peripheral blood lymphocytes and cloned lymphocyte lines. This repressive effect on the HTLV-I LTR by the HCMV IE-2 gene product(s) represent a unique interaction between two viruses capable of infecting the same target cell in vivo. Such an interaction may have important implications for disease expression associated with HTLV-I infection.