Human JC virus nuclear factor 1 binding motifs and large tumor antigen region required for transactivation of late promoter

Kotlo U. Kumar, Laxminarayana R. Devireddy, Shou Ching Tang, Alan Pater, Mary M. Pater

Research output: Contribution to journalArticle

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Abstract

The nuclear factor 1 (NF-1) motifs, NF-1 II/III, in the two 98-bp repeats of the transcription-regulatory region of JC virus (JCV), have a critical role in brain-specific transcription from the JCV early promoter- enhancer. In this study, the role of these motifs in transactivation of the JCV late promoter-enhancer (JCVL) was examined in differentiating glial P19 embryonal carcinoma cells. The expression of papovaviral large tumor antigen (T-Ag) in the glial cells was shown by double immunofluorescence assays. By using site-directed mutagenesis and in vivo assays, the two wild-type NF-1 II/III sites, but not the third site, were found to be essential for the transactivation of JCV(L) by JCV T-Ag. In vitro transcription assays confirmed this specific transactivation and the transactivation was abolished by T-Ag antibody. In electrophoretic mobility shift assays, expression of JCV T-Ag increased the binding of a factor(s) to the 98-bp repeat. T-Ag antibody abolished the increase of binding. Binding assays with oligonucleotides of NF-1 II/III motifs showed that the increased binding specifically required the wild-type NF-1 II/III sequences and confirmed the requirement of T-Ag. To determine the region of T-Ag necessary for transactivation of JCV(L), the coding sequences were mutated. The amino-terminal region of JCV Ag in amino acids 1-437 was essentially required for efficient transactivation. These results indicated that transactivation of JCV(L) and increased binding require a factor(s) found specifically in glial cells, the JCV NF-1 II/III sites, and the T-Ag amino-terminal region.

Original languageEnglish (US)
Pages (from-to)473-481
Number of pages9
JournalJournal of Neurochemistry
Volume67
Issue number2
StatePublished - Aug 1 1996

Fingerprint

NFI Transcription Factors
JC Virus
Neoplasm Antigens
Viruses
Transcriptional Activation
Assays
Transcription
Neuroglia
Neoplasm Antibodies
Embryonal Carcinoma Stem Cells
Electrophoretic mobility
Mutagenesis
Antibodies
Nucleic Acid Regulatory Sequences
Electrophoretic Mobility Shift Assay
Site-Directed Mutagenesis
Oligonucleotides
Fluorescent Antibody Technique
Brain
Cells

Keywords

  • JC virus
  • Late promoter
  • Nuclear factor 1 sites
  • T-Ag
  • Transactivation

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

Cite this

Kumar, K. U., Devireddy, L. R., Tang, S. C., Pater, A., & Pater, M. M. (1996). Human JC virus nuclear factor 1 binding motifs and large tumor antigen region required for transactivation of late promoter. Journal of Neurochemistry, 67(2), 473-481.

Human JC virus nuclear factor 1 binding motifs and large tumor antigen region required for transactivation of late promoter. / Kumar, Kotlo U.; Devireddy, Laxminarayana R.; Tang, Shou Ching; Pater, Alan; Pater, Mary M.

In: Journal of Neurochemistry, Vol. 67, No. 2, 01.08.1996, p. 473-481.

Research output: Contribution to journalArticle

Kumar, Kotlo U. ; Devireddy, Laxminarayana R. ; Tang, Shou Ching ; Pater, Alan ; Pater, Mary M. / Human JC virus nuclear factor 1 binding motifs and large tumor antigen region required for transactivation of late promoter. In: Journal of Neurochemistry. 1996 ; Vol. 67, No. 2. pp. 473-481.
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AU - Pater, Alan

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AB - The nuclear factor 1 (NF-1) motifs, NF-1 II/III, in the two 98-bp repeats of the transcription-regulatory region of JC virus (JCV), have a critical role in brain-specific transcription from the JCV early promoter- enhancer. In this study, the role of these motifs in transactivation of the JCV late promoter-enhancer (JCVL) was examined in differentiating glial P19 embryonal carcinoma cells. The expression of papovaviral large tumor antigen (T-Ag) in the glial cells was shown by double immunofluorescence assays. By using site-directed mutagenesis and in vivo assays, the two wild-type NF-1 II/III sites, but not the third site, were found to be essential for the transactivation of JCV(L) by JCV T-Ag. In vitro transcription assays confirmed this specific transactivation and the transactivation was abolished by T-Ag antibody. In electrophoretic mobility shift assays, expression of JCV T-Ag increased the binding of a factor(s) to the 98-bp repeat. T-Ag antibody abolished the increase of binding. Binding assays with oligonucleotides of NF-1 II/III motifs showed that the increased binding specifically required the wild-type NF-1 II/III sequences and confirmed the requirement of T-Ag. To determine the region of T-Ag necessary for transactivation of JCV(L), the coding sequences were mutated. The amino-terminal region of JCV Ag in amino acids 1-437 was essentially required for efficient transactivation. These results indicated that transactivation of JCV(L) and increased binding require a factor(s) found specifically in glial cells, the JCV NF-1 II/III sites, and the T-Ag amino-terminal region.

KW - JC virus

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