Infection and replication of Tat- @entity1 XXXX viruses: genetic analyses of LTR and tat mutations in primary and long-term @entity1 lymphoid cells.

@entity1 @entity465 @entity645 @entity2858
Tat is an essential regulatory protein for the replication of @entity1 @entity465 virus (HIV). Mutations in the tat gene have been shown to block @entity645 replication in @entity1 T cells. Several studies have established that Tat releases an elongation block to the transcription of @entity645 long terminal repeat (LTR); however, it is not known whether this mechanism alone is sufficient to explain the block to @entity645 replication in @entity1 T cells when Tat is absent. It is possible that Tat is also needed for other functions during @entity645 replication. To test these hypotheses, we studied several tat mutants, including two stop codon mutants and one deletion mutant using replication-competent @entity2858 constructs carrying wild-type or mutant LTRs with modifications in the NF-kappa B and/or Sp1 binding sites. In this study, we show that Tat- @entity2858 with wild-type LTRs can replicate in HeLa cells, and the virus produced from HeLa cells can infect primary peripheral blood lymphocytes and macrophages. It was found that the propagation of the Tat mutants containing wild-type LTRs was less efficient than that of the LTR-modified Tat mutants. Large amounts of viral RNA and particles were synthesized in infections established using the tat mutants that contain modified LTRs. However, this efficient propagation of the LTR-modified tat mutants was restricted to some lymphoid cell lines that have been transformed with other viruses. Thus, despite its essential role for releasing an elongation block, Tat is not otherwise absolutely required for synthesis of full-length @entity645 transcripts and assembly of virus particles. Direct sequencing of the viral genomes and reinfection kinetics showed no evidence of wild-type reversion even after prolonged infection with the Tat- virus. The implications for in vivo @entity2858 replication and potential application of this system to the study of alternative Tat function are discussed.
