Identification of limiting steps for efficient trans-activation of HIV-1 promoter by Tat in Saccharomyces cerevisiae.
Cellular context is an important determinant for the activity of Tat, the trans-activator of human immunodeficiency virus (HIV). We have investigated HIV-1 promoter expression and trans-activation in Saccharomyces cerevisiae to provide clues about the limiting steps for Tat activity in this organism. A minimal 43-nucleotide HIV promoter (HIV43) has the activity of a weak yeast promoter in the presence or absence of various enhancer binding sites (bs), whereas the entire long terminal repeat is not expressed. None of these constructs could be trans-activated by Tat. Fusion proteins Gal4 binding domain (BD)-Tat48 and Gal4BD-Tat72 are active with different efficiencies on various yeast promoters that have Gal4 bs. They have 70 and 50% of Gal4 wild type activity on hybrid HIV promoters fused to Gal4 bs only in the presence of AP1 bs. This study shows that trans-activation of the HIV-1 promoter by Tat occurs in yeast when Tat is targeted to the promoter and a functional enhancer activity is present. Sp1 function and Tat transfer from the RNA to the promoter are two major elements for in vivo trans-activation of HIV-1 that are defective in S. cerevisiae but can be replaced by functional equivalents.
Daviet
L
L
Unité 332, INSERM, Institut Cochin de Génétique Moléculaire, 22, rue Méchain, 75014 Paris, France.
Bois
F
F
Battisti
P L
PL
Gatignol
A
A
D001483:Base Sequence; D004261:DNA Replication; D004268:DNA-Binding Proteins; D004742:Enhancer Elements, Genetic; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000378:metabolism*; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015497:HIV-1 / Q000235:genetics*; D008969:Molecular Sequence Data; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010692:Phleomycins / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000821:virology; D029701:Saccharomyces cerevisiae Proteins; D016329:Sp1 Transcription Factor / Q000378:metabolism; D013045:Species Specificity; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D016662:Virus Integration; D014779:Virus Replication; D054322:tat Gene Products, Human Immunodeficiency Virus
D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; D015696:Gene Products, tat; D019384:Nucleic Acid Synthesis Inhibitors; D010692:Phleomycins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus