RESUMO
Strategies to inhibit the spread of HIV infection consist of a number of specific molecular approaches. Since viral production is dependent upon Tat-mediated transactivation of the HIV promoter through the Tat activating region (TAR), tat antisense RNA, anti-tat ribozymes, TAR decoys and dominant negative Tat mutant proteins have been suggested as therapeutic inhibitors. We produced and tested several Tat mutant proteins, including a newly generated form Tat delta 58, for the ability to inhibit Tat-mediated transactivation and HIV production. In addition, we generated a new Tat fusion mutant between a C-terminus truncated form of Tat (Tat delta 53) and the Drosophila Engrailed (Eng) transcription repressor domain to test the hypothesis that transcriptional repression can be targeted to the HIV promoter. This fusion mutant was also examined for its capacity to block both Tat-mediated transactivation and HIV replication. We show that three mutants Tat delta 53. Tat delta 58 and Tat delta 53/Eng result in a transdominant phenotype inhibiting wild-type Tat-mediated transactivation, and that the inhibiting potential is increased by the presence of the entire basic domain or the fusion of a repressor domain. However, only the transdominant mutants Tat delta 58 and Tat delta 53/Eng significantly inhibit HIV-1 replication after infection of transfected T cell lines. These results demonstrate the potent inhibiting activity of Tat mutants on HIV replication, and suggest a synergistic effect of Tat transdominant mutant fusion with the Drosophila Engrailed transcription repressor domain.
Assuntos
Produtos do Gene tat/genética , Terapia Genética/métodos , Infecções por HIV/prevenção & controle , HIV-1/fisiologia , Ativação Transcricional/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Linhagem Celular , Relação Dose-Resposta a Droga , Proteínas de Drosophila , Repetição Terminal Longa de HIV/genética , Proteínas de Homeodomínio/genética , Humanos , Mutação , Proteínas Recombinantes de Fusão/farmacologia , Linfócitos T , Fatores de Transcrição/genética , Produtos do Gene tat do Vírus da Imunodeficiência HumanaRESUMO
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