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Journal of virology2004; 78(7); 3407-3418; doi: 10.1128/jvi.78.7.3407-3418.2004

PU.1 binding to ets motifs within the equine infectious anemia virus long terminal repeat (LTR) enhancer: regulation of LTR activity and virus replication in macrophages.

Abstract: Binding of the transcription factor PU.1 to its DNA binding motif regulates the expression of a number of B-cell- and myeloid-specific genes. The long terminal repeat (LTR) of macrophage-tropic strains of equine infectious anemia virus (EIAV) contains three PU.1 binding sites, namely an invariant promoter-proximal site as well as two upstream sites. We have previously shown that these sites are important for EIAV LTR activity in primary macrophages (W. Maury, J. Virol. 68:6270-6279, 1994). Since the sequences present in these three binding motifs are not identical, we sought to determine the role of these three sites in EIAV LTR activity. While DNase I footprinting studies indicated that all three sites within the enhancer were bound by recombinant PU.1, reporter gene assays demonstrated that the middle motif was most important for basal levels of LTR activity in macrophages and that the 5' motif had little impact. The impact of the 3' site became evident in Tat transactivation studies, in which the loss of the site reduced Tat-transactivated expression 40-fold. In contrast, elimination of the 5' site had no effect on Tat-mediated activity. Binding studies were performed to determine whether differences in PU.1 binding affinity for the three sites correlated with the relative impact of each site on LTR transcription. While small differences were observed in the binding affinities of the three sites, with the promoter-proximal site having the strongest binding affinity, these differences could not account for the dramatic differences observed in the transcriptional effects. Instead, the promoter-proximal position of the 3' motif appeared to be critical for its transcriptional impact and suggested that the PU.1 sites may serve different roles depending upon the location of the sites within the enhancer. Infectivity studies demonstrated that an LTR containing an enhancer composed of the three PU.1 sites was not sufficient to drive viral replication in macrophages. These findings indicate that while the promoter-proximal PU.1 site is the most critical site for EIAV LTR activity in the presence of Tat, other elements within the enhancer are needed for EIAV replication in macrophages.
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Publication Date: 2004-03-16 PubMed ID: 15016863PubMed Central: PMC371083DOI: 10.1128/jvi.78.7.3407-3418.2004Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research examines how the transcription factor PU.1 binds to different motifs in the replication process of equine infectious anemia virus (EIAV) and how it impacts the virus’ ability to activate certain genes. Their findings highlight the complexity and significance of these binding processes in the functioning and control of the virus.

Understanding PU.1 and EIAV

  • PU.1 is a transcription factor that helps control gene expression, especially in B-cells and myeloid cells.
  • EIAV (equine infectious anemia virus) is a horse disease that can help us understand similar conditions in humans. It uses the long terminal repeat (LTR) as a virus promoter, which contains three PU.1 binding sites.
  • Previous research found these sites crucial for EIAV LTR activity in macrophages, types of white blood cells that play key roles in immunity.

The Study’s Methods and Findings

  • The researchers evaluated the roles of the three PU.1 binding sites and their contribution to EIAV LTR activity. Their experiments included DNase I footprinting studies and reporter gene assays.
  • The middle motif seemed the most important for maintaining basal levels of LTR activity in macrophages, while the 5′ motif impact was minor.
  • In the study, they also focused on the effect of Tat, a protein that helps the virus replicate. Removing the 3′ site led to a significant reduction in Tat-transactivated expression, however, the 5′ site’s elimination did not affect this activity.

Observations about PU.1 Binding Affinity

  • They attempted to understand whether variations in PU.1 binding affinity for the three sites correlated with each site’s impact on LTR transcription. They noted slight variations in binding affinities, but these could not explain the significant differences in transcriptional impact.
  • Notably, the position of the 3′ site, closest to the promoter, was instrumental for its impact, suggesting that the location of PU.1 sites within the enhancer can determine their roles.

Infectivity Studies

  • The researchers also conducted infectivity studies and found that an LTR with an enhancer made of the three PU.1 sites was insufficient to drive viral replication in macrophages.
  • This highlights that, despite the promoter-proximal PU.1 site being crucial for EIAV LTR activity in the presence of Tat, other elements within the enhancer are necessary for EIAV replication.

Cite This Article

APA
Hines R, Sorensen BR, Shea MA, Maury W. (2004). PU.1 binding to ets motifs within the equine infectious anemia virus long terminal repeat (LTR) enhancer: regulation of LTR activity and virus replication in macrophages. J Virol, 78(7), 3407-3418. https://doi.org/10.1128/jvi.78.7.3407-3418.2004

Publication

Journal of virology
ISSN: 0022-538X
NlmUniqueID: 0113724
Country: United States
Language: English
Volume: 78
Issue: 7
Pages: 3407-3418

Researcher Affiliations

Hines, Robert
  • Division of Basic Biomedical Science, University of South Dakota, Vermillion, South Dakota 57069, USA.
Sorensen, Brenda R
    Shea, Madeline A
      Maury, Wendy

        MeSH Terms

        • Base Sequence
        • Binding Sites
        • Cell Line
        • DNA Footprinting
        • DNA-Binding Proteins / chemistry
        • DNA-Binding Proteins / genetics
        • DNA-Binding Proteins / metabolism
        • Enhancer Elements, Genetic / genetics
        • Gene Expression Regulation, Viral
        • HIV Enhancer / genetics
        • Humans
        • Infectious Anemia Virus, Equine / genetics
        • Macrophages / virology
        • Models, Molecular
        • Molecular Sequence Data
        • Mutation / genetics
        • Nucleic Acid Conformation
        • Promoter Regions, Genetic / genetics
        • Protein Binding
        • Protein Structure, Tertiary
        • Proto-Oncogene Proteins / chemistry
        • Proto-Oncogene Proteins / genetics
        • Proto-Oncogene Proteins / metabolism
        • Proto-Oncogene Proteins c-ets
        • Response Elements / genetics
        • Terminal Repeat Sequences / genetics
        • Trans-Activators / chemistry
        • Trans-Activators / genetics
        • Trans-Activators / metabolism
        • Transcription Factors / metabolism
        • Virus Replication

        Grant Funding

        • CA72063 / NCI NIH HHS

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        Citations

        This article has been cited 6 times.
        1. Wang XF, Liu Q, Wang YH, Wang S, Chen J, Lin YZ, Ma J, Zhou JH, Wang X. Characterization of Equine Infectious Anemia Virus Long Terminal Repeat Quasispecies In Vitro and In Vivo. J Virol 2018 Apr 15;92(8).
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