title: Negative-strand RNA transcripts are produced in human immunodeficiency virus type 1-infected cells and patients by a novel promoter downregulated by Tat.
abstract: Current understanding of human immunodeficiency virus type 1 (HIV-1) transcription is based on unidirectional expression of transcripts with positive-strand polarity from the 5' long terminal repeat. We now report HIV-1 transcripts with negative-strand polarity obtained from acutely and chronically infected cell lines by use of a template orientation-specific reverse transcriptase-PCR assay. These findings were confirmed in natural infection by analysis of RNA derived from peripheral blood mononuclear cell samples from 15 HIV-1-infected patients. A cDNA derived from a 2.3-kb polyadenylated HIV-1 RNA with negative-strand polarity which encodes a highly conserved 189-amino-acid open reading frame antiparallel to the envelope gene was isolated from acutely infected A3.01 cells. Through use of reporter gene constructions, we further found that a novel negative-strand promoter functions within the negative response element of the 3' long terminal repeat, which is downregulated by coexpression of Tat. Site-directed mutagenesis experiments demonstrated that NF-kappa B I and USF sites are crucial for negative-strand promoter activity. These data extend the coding capacity of HIV-1 and suggest a role for antisense regulation of the viral life cycle.
authors.affiliation: Department of Retroviral Research, Walter Reed Army Institute of Research, Rockville, Maryland.
authors.forename: N L
authors.initials: NL
authors.lastname: Michael
authors.forename: M T
authors.initials: MT
authors.lastname: Vahey
authors.forename: L
authors.initials: L
authors.lastname: d'Arcy
authors.forename: P K
authors.initials: PK
authors.lastname: Ehrenberg
authors.forename: J D
authors.initials: JD
authors.lastname: Mosca
authors.forename: J
authors.initials: J
authors.lastname: Rappaport
authors.forename: R R
authors.initials: RR
authors.lastname: Redfield
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002478:Cells, Cultured; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000235:genetics; D015536:Down-Regulation; D015870:Gene Expression; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D015696:Gene Products, tat / Q000494:pharmacology*; D015658:HIV Infections / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016328:NF-kappa B; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis; D012367:RNA, Viral / Q000096:biosynthesis; D017384:Sequence Deletion; D014158:Transcription, Genetic / Q000187:drug effects*; D054322:tat Gene Products, Human Immunodeficiency Virus
chemical_list: D018076:DNA, Complementary; D015696:Gene Products, tat; D016328:NF-kappa B; D012333:RNA, Messenger; D012367:RNA, Viral; D054322:tat Gene Products, Human Immunodeficiency Virus