ESSENTIAL ROLE OF NF-KAPPA-B IN TRANSACTIVATION OF THE HUMAN-IMMUNODEFICIENCY-VIRUS LONG TERMINAL REPEAT BY THE HUMAN CYTOMEGALOVIRUS IE1 PROTEIN

The 72 kDa IE1 protein of human cytomegalovirus (HCMV) is one of a few viral regulatory proteins expressed immediately after infection of a host cell. Although it is now well-established that IE1 is a potent tr anscriptional activator of the human immunodeficiency virus (HIV) long terminal repeat (LTR), the identity of the nucleotide sequence respon sive to IE1 remains elusive and the molecular mechanism of this intera ction is not well-understood. We have constructed various LTR mutants and tested them for their ability to be activated by IE1 using transie nt transfection assays. Mutations in the NF-kappa B sites, of either a few changes in the nucleotide sequence or a deletion of the entire re gion, abrogated IE1-driven transactivation. Deletion of the Tat-respon sive element (TAR) had no significant effect on reporter expression. M utations in the Sp1 sites or the TATA box significantly lowered LTR ac tivity, but this is probably due to an effect on the general transcrip tion system, as these elements are also required for the transactivati on of the LTR by many stimulators including Tat, tumour necrosis facto r alpha (TNF-alpha), E1A/E1B and phorbol myristate acetate (PMA). In a ddition, gel retardation analysis demonstrated that NF-kappa B activit y was significantly increased in human T lymphoid H9 and monocytic U93 7 cell lines constitutively expressing IE1. Taken together, these data suggest that NF-kappa B plays a central role in the IE1 transactivati on of the HIV LTR.