Journal article
A highly conserved HIV-1 clade B downstream C/EBP binding site affects C/EBP and Tat transactivation
Journal of neurovirology, Vol.12
01 May 2006
Abstract
Previous studies have shown that at least one CCAAT enhancer binding protein (C/EBP) site upstream of the TATA box and full-length C/EBP beta protein is necessary for human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) activity in cells of the monocyte/macrophage lineage. However, no studies have been performed concerning C/EBP sites downstream (DS) of the start of transcription. Analyses of 115 clade B LTRs have indicated that there are three potential C/EBP sites within the downstream LTR. Electrophoretic mobility shift (EMS) analyses demonstrated that only one of the three sites (DS3, +158 to +172) was able to bind members of the C/EBP family. Analyses of 115 clade C, 21 clade A and 19 clade D LTRs indicated that this downstream site is highly conserved among different HIV-1 clades, suggesting the presence of a functionally important cis-acting element. EMS analysis of naturally occurring variants within DS3 demonstrated altered C/EBP recruitment. To perform functional studies, the 6C variant was selected (G-to-C change at position 6) because it resulted in a knockout phenotype without creating an alternate binding site. Results from transfection studies utilizing the parental wild type LAI LTR (LTR-WT) containing the knock-out 6C variant (LTR-DS3-6C) demonstrated loss of binding to this site results in an approximate 50% decrease in basal LTR activity in the U-937 monocytic cell line. Full-length C/EBP beta induction of LTR-WT was reduced 3-fold by loss of C/EBP binding to DS3. Interestingly, Tat induction was decreased 4-fold, suggesting that an alteration in the structural integrity of this site may perturbate the interaction of Tat with the neighboring TAR element that may involve C/EBP/Tat interactions. Co-transfection assays demonstrated that full-length C/EBP synergistically enhanced Tat transcriptional activity with both LTR-WT and LTR-DS3-6C, with a higher induction level in LTR-WT. These observations suggest the highly conserved DS3 site is important for HIV-1 LTR transcription and may be a critical element required for HIV-1 activation.
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Details
- Title
- A highly conserved HIV-1 clade B downstream C/EBP binding site affects C/EBP and Tat transactivation
- Creators
- Yujie LiuM NonnemacherE KilareskiA AlexakiB Wigdahl
- Publication Details
- Journal of neurovirology, Vol.12
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Microbiology and Immunology
- Identifiers
- 991019170338304721