The transactivation induced by human herpesvirus 6 (HHV-6) on the HIV-1 promoter was studied both in the presence and in the absence of the retroviral transactivator protein (Tat) constitutively expressed in Jurkat cells. Jurkat-tatcells were infected with HHV-6, transfected with a plasmid containing HIV-1 long terminal repeat (LTR)/chloramphenicol acetyltransferase (CAT), and CAT assays were performed. HHV-6 infection in the presence of Tat resulted in significantly higher LTR activation than that observed by Tat or HHV-6 alone, indicating that HHV-6 and Tat interact synergically. Furthermore, it was demonstrated that expression of HIV-1tatinhibits HHV-6 replication, as shown by a 3.6–15.4-fold reduction in infectious yield. We suggest that HHV-6 could trigger HIV reactivation in HIV-seropositive patients which, in turn, could inhibit HHV-6 production.