SEQUENCE ELEMENTS DOWNSTREAM OF THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 LONG TERMINAL REPEAT ARE REQUIRED FOR EFFICIENT VIRAL GENE-TRANSCRIPTION

We investigated the role of a 54-nucleotide region (+200 to +253) loca ted downstream of the HIV-1 long terminal repeat (LTR) on virus gene e xpression and found, using RT-PCR and p24 CA analysis, that deletion o f this region inhibited synthesis of both viral RNA and protein. CAT a ssays showed that these results were attributable to decreased transcr iption efficiency of the HIV-1 LTR and not to the stability of the RNA transcripts produced. Further deletional analysis and transfection st udies showed that the most important sequences with regard to proviral DNA expression were located between nucleotide positions +218 and +23 7. Furthermore, substitutional mutational analysis showed that a CTCTC TC sequence at positions +227 to +233, homologous to the pyrimidine-ri ch initiator (Inr) region found in several promoters, was required for efficient production of both viral RNA and protein. Deletion of the s equence +200 to +217, homologous to the interferon-stimulated response element (ISRE), resulted in impaired LTR promoter activity and decrea sed synthesis of viral RNA and protein. However, when the latter regio n was replaced by homologous ISRE sequences from an interferon-stimula ted gene (ISG-54), an even more severe effect on HIV gene expression a nd replication was observed, suggesting that ISRE-like sequences in HI V act differently from homologous sequences in interferon-responsive c ellular genes. (C) 1997 Academic Press Limited.