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Journal of Virology, February 1999, p. 1331-1340, Vol. 73, No. 2
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Evolution of the Human Immunodeficiency Virus Type 1 Long Terminal Repeat Promoter by Conversion of an NF-kappa B Enhancer Element into a GABP Binding Site

Koen Verhoef,1 Rogier W. Sanders,1 Veronique Fontaine,2 Shigetaka Kitajima,3 and Ben Berkhout1,*

Department of Human Retrovirology1 and Department of Medical Microbiology,2 Academic Medical Center, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands, and Department of Biochemical Genetics, Medical Research Institute, Tokyo Medical and Dental University, Bunkyou-ku 113, Tokyo, Japan3

Received 30 July 1998/Accepted 30 October 1998

Human immunodeficiency virus type 1 (HIV-1) transcription is regulated by the viral Tat protein and cellular factors, of which the concentration and activity may depend on the cell type. Viral long terminal repeat (LTR) promoter sequences are therefore optimized to suit the specific nuclear environment of the target host cell. In long-term cultures of a Tat-defective, poorly replicating HIV-1 mutant, we selected for a faster-replicating virus with a 1-nucleotide deletion in the upstream copy of two highly conserved NF-kappa B binding sites. The variant enhancer sequence demonstrated a severe loss of NF-kappa B binding in protein binding assays. Interestingly, we observed a new binding activity that is specific for the variant NF-kappa B sequence and is present in the nuclear extract of unstimulated cells that lack NF-kappa B. These results suggest that inactivation of the NF-kappa B site coincides with binding of another transcription factor. Fine mapping of the sequence requirements for binding of this factor revealed a core sequence similar to that of Ets binding sites, and supershift assays with antibodies demonstrated the involvement of the GABP transcription factor. Transient transfection experiments with LTR-chloramphenicol acetyltransferase constructs indicated that the variant LTR promoter is specifically inhibited by GABP in the absence of Tat, but this promoter was dramatically more responsive to Tat than the wild-type LTR. Introduction of this GABP site into the LAI virus yielded a specific gain of fitness in SupT1 cells, which contain little NF-kappa B protein. These results suggest that GABP potentiates Tat-mediated activation of LTR transcription and viral replication in some cell types. Conversion of an NF-kappa B into a GABP binding site is likely to have occurred also during the worldwide spread of HIV-1, as we noticed the same LTR modification in subtype E isolates from Thailand. This typical LTR promoter configuration may provide these viruses with unique biological properties.

* Corresponding author. Mailing address: Department of Human Retrovirology, Academic Medical Center, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands. Phone: 31-20-5664822. Fax: 31-20-6916531. E-mail: b.berkhout{at}amc.uva.nl.

Journal of Virology, February 1999, p. 1331-1340, Vol. 73, No. 2
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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