992. Suppression of the HIV-1 Long Terminal Repeat by Transducible Artificial Transcription Factors

Abstract

Top of pageAbstract The long terminal repeats (LTR) found in the genomes of lentiviruses, including the human immunodeficiency virus (HIV), play a crucial role in viral pathogenesis. The LTRs contain DNA elements that interact with various types of transcriptional regulatory proteins found in host cells; these include the basal transcription machinery, cell-type-and, gene-specific transcription factors found in plurality of cells. Using human zinc finger domains, we designed artificial transcription factors that target the Sp-1 binding sites and TATA box of the HIV-1 type B LTR. In transiently-transfected Hela cells, these transcription factors strongly suppressed LTR-directed transcription of a reporter gene, which is tightly controlled by co-expression of the HIV-Tat protein. In contrast to previous findings by Reynolds et al. (PNAS, 2003, 100(4):1615-1620) these artificial zinc finger transcription factors inhibited HIV-1 LTR activity whether or not the zinc finger domains were fused to the transcriptional repression domain such as KRAB. We next sought to determine whether LTR activity in Hela cells transiently transfected with reporter gene could be inhibited by artificial transcription factors delivered to the cells in the form of protein. Rapid delivery of such proteins into white blood cells may be more efficacious than gene delivery, such as that mediated by frequently used viral vectors. To this end, we expressed artificial zinc finger transcription factors that were fused to the PTD (protein transduction domain) of the HIV-1 Tat protein. The resulting recombinant transcription factors were successfully delivered to virtually all cells in culture and repressed LTR-directed transcriptional activation of a reporter gene. We finally confirmed that the LTR suppression by transducible artificial transcription factors strongly inhibited HIV-1 replication from cells infected with a replication-competent HIV-1 strain. Our results strongly suggest that transducible artificial transcription factors that inhibit LTR activity represent potential drugs for the inhibition of HIV-1 replication|[mdash]|a hallmark of AIDS pathogenesis.