title: The HIV-1 Tat protein has a versatile role in activating viral transcription.
abstract: It is generally acknowledged that the Tat protein has a pivotal role in HIV-1 replication because it stimulates transcription from the viral long terminal repeat (LTR) promoter by binding to the TAR hairpin in the nascent RNA transcript. However, a multitude of additional Tat functions have been suggested. The importance of these functions is difficult to assess in replication studies with Tat-mutated HIV-1 variants because of the dominant negative effect on viral gene expression. We therefore used an HIV-1 construct that does not depend on the Tat-TAR interaction for transcription to reevaluate whether or not Tat has a second essential function in HIV-1 replication. This HIV-rtTA variant uses the incorporated Tet-On gene expression system for activation of transcription and replicates efficiently upon complete TAR deletion. Here we demonstrated that Tat inactivation does nevertheless severely inhibit replication. Upon long-term culturing, the Tat-minus HIV-rtTA variant acquired mutations in the U3 region that improved promoter activity and reestablished replication. We showed that in the absence of a functional TAR, Tat remains important for viral transcription via Sp1 sequence elements in the U3 promoter region. Substitution of these U3 sequences with nonrelated promoter elements created a virus that replicates efficiently without Tat in SupT1 T cells. These results indicate that Tat has a versatile role in transcription via TAR and U3 elements. The results also imply that Tat has no other essential function in viral replication in cultured T cells.
authors.affiliation: Laboratory of Experimental Virology, Academic Medical Center, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands. a.t.das@amc.uva.nl
authors.forename: Atze T
authors.initials: AT
authors.lastname: Das
authors.forename: Alex
authors.initials: A
authors.lastname: Harwig
authors.forename: Ben
authors.initials: B
authors.lastname: Berkhout
mesh_terms: D002460:Cell Line; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D011401:Promoter Regions, Genetic; D012367:RNA, Viral / Q000235:genetics; D014158:Transcription, Genetic; D014779:Virus Replication; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*
chemical_list: D012367:RNA, Viral; D054322:tat Gene Products, Human Immunodeficiency Virus