AEGiS-13IAC: Repression of HIV-1 replication by the transcrans-criptional repressor protein NFAT-KRAB as a model for gene therapy.

13th International AIDS Conference

Durban, South Africa - July 9-July 14, 2000

Repression of HIV-1 replication by the transcrans-criptional repressor protein NFAT-KRAB as a model for gene therapy.

Int Conf AIDS 2000 Jul 9-14; 13:(abstract no. TuOrA406)

Schneider J, Hahne J
J. Schneider, University of Freiburg, Department of Virology, Hermann-Herder-Str. 11, D-79104 Freiburg, Germany, Tel.: +49 761 203 65 88, Fax: +49 761 203 66 39, E-mail: schf@ukl.uni-freiburg.de

BACKGROUND: The initiation of HIV-transcription is regulated by cellular proteins which bind to the enhancer region of the integrated provirus. Therefore recombinant proteins consisting of enhancer-binding and transcription repression domains can be envisaged as efficient intracellular repressors of HIV. In our model, the DNA binding domain of NFAT (Nuclear Factor of Activated T-cells) and the Krnppel-associated Box (KRAB) domain of KOX-1 were fused to a HIV targetted novel repressor protein. To minimize imunological elimination of the transfected cells in vivo both domains are of human origin.

METHODS: An expression plasmid for NFAT-KRAB was constructed and consistently NFAT-KRAB expressing clones were selected upon transfection of Jurkat cells. In addition, primary lymphocytes and macrophages were transiently transfected with the NFAT-KRAB expression plasmid. The repressive effect of NFAT-KRAB was evaluated by measuring virus production in HIV-1IIIB-, or HIV-1BAL infected cultures by a commercial p24-antigen ELISA.

RESULTS: Virus production was suppressed by the NFAT-KRAB protein in the Jurkat cell clones and in primary cells. The specificity of suppression was corroborated by its reversion with the NFAT-inactivating drug Cyclosporin A. Expression of NFAT-KRAB was compatible with normal growth of Jurkat-NFAT-KRAB cells and the protective effect of NFAT-KRAB was revealed as increased cell survial after HIV-infection in transfected versus untransfected cultures.

CONCLUSIONS: The chimeric NFAT-KRAB reduces production of two different HIV isolates and the HIV-induced cytopathic effect. Therefore the NFAT-KRAB repressor protein appears to be suited for intracellular immunization against HIV-1.

Keywords: AEGIS, Repressor Proteins, HIV-1, Virus Replication, Transcription Factors, DNA-Binding Proteins, Gene Therapy, Transcription, Genetic, Transfection, HIV Infections, Jurkat Cells, Plasmids, T-Lymphocytes, transcription factor NF-AT, Human, virology, genetics
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