Reconstitution of papillomavirus E2-mediated plasmid maintenance in Saccharomyces cerevisiae by the Brd4 bromodomain protein

Angela R. Brannon, Julia A. Maresca, Jef D. Boeke, Munira A. Basrai, Alison A. McBride

Research output: Contribution to journalArticle

Abstract

The papillomavirus E2 protein functions in viral transcriptional regulation, DNA replication, and episomal genome maintenance. Viral genomes are maintained in dividing cells by attachment to mitotic chromosomes by means of the E2 protein. To investigate the chromosomal tethering function of E2, plasmid stability assays were developed in Saccharomyces cerevisiae to determine whether the E2 protein could maintain plasmids containing the yeast autonomous replication sequence replication element but with the centromeric element replaced by E2-binding sites. E2 expression was not sufficient to maintain such plasmids, but plasmid stability could be rescued by expression of the mammalian protein Brd4. In the presence of both Brd4 and E2 proteins, plasmids with multiple E2-binding sites were stable without selection. S. cerevisiae encodes a homolog of Brd4 named Bdf1 that does not contain the C-terminal domain that interacts with the E2 protein. A fusion protein of Bdf1 and the Brd4 C-terminal "tail" could support E2-mediated plasmid maintenance in yeast. Using a panel of mutated E2 proteins, we determined that plasmid stability required the ability of E2 to bind DNA and to interact with Brd4 and mammalian mitotic chromosomes but did not require its replication initiation and transactivation functions. The S. cerevisiae-based plasmid maintenance assays described here are invaluable tools for dissecting mechanisms of episomal viral genome replication and screening for additional host protein factors involved in plasmid maintenance.

Original languageEnglish (US)
Pages (from-to)2998-3003
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number8
DOIs
StatePublished - Feb 22 2005

Fingerprint

Saccharomyces cerevisiae
Plasmids
Maintenance
Proteins
Viral Genome
DNA Replication
Integration Host Factors
Yeasts
Mammalian Chromosomes
Binding Sites
Transcriptional Activation
Chromosomes
Genome
DNA

Keywords

  • Bovine papillomavirus
  • Human papillomavirus
  • Replication
  • Segregation
  • Yeast

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Reconstitution of papillomavirus E2-mediated plasmid maintenance in Saccharomyces cerevisiae by the Brd4 bromodomain protein. / Brannon, Angela R.; Maresca, Julia A.; Boeke, Jef D.; Basrai, Munira A.; McBride, Alison A.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 102, No. 8, 22.02.2005, p. 2998-3003.

Research output: Contribution to journalArticle

Brannon, Angela R. ; Maresca, Julia A. ; Boeke, Jef D. ; Basrai, Munira A. ; McBride, Alison A. / Reconstitution of papillomavirus E2-mediated plasmid maintenance in Saccharomyces cerevisiae by the Brd4 bromodomain protein. In: Proceedings of the National Academy of Sciences of the United States of America. 2005 ; Vol. 102, No. 8. pp. 2998-3003.
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