Genome wide, supercoiling-dependent in vivo binding of a viral protein involved in DNA replication and transcriptional control

Víctor González-Huici, Margarita Salas, José M Hermoso

Research output: Contribution to journalArticlepeer-review

Abstract

Protein p6 of Bacillus subtilis bacteriophage Phi29 is essential for phage development. In vitro it activates the initiation of DNA replication and is involved in the early to late transcriptional switch. These activities require the formation of a nucleoprotein complex in which the DNA forms a right-handed superhelix wrapping around a multimeric protein core. However, there was no evidence of p6 binding to Phi29 DNA in vivo. By crosslinking, chromatin immunoprecipitation and real-time PCR we show that protein p6 binds to most, if not all, the viral genome in vivo, although with higher affinity for both DNA ends, which contain the replication origins. In contrast, the affinity for plasmid DNA is negligible, but greatly increases when the negative supercoiling decreases, as shown in vivo by treatment of cells with novobiocin and in vitro by fluorescence quenching with plasmids with different topology. In conclusion, binding of protein p6 all along the Phi29 genome strongly suggests that its functions in replication and transcription control could be local outcomes of a more global role as a histone-like protein. The p6 binding dependence on DNA topology could explain its preferential binding to viral with respect to bacterial DNA, whose level of negative supercoiling is presumably higher than that of Phi29 DNA.

Original languageEnglish
Pages (from-to)2306-14
Number of pages9
JournalNucleic Acids Research
Volume32
Issue number8
DOIs
Publication statusPublished - 2004

Keywords

  • Bacillus Phages
  • DNA, Superhelical
  • DNA, Viral
  • Gene Expression Regulation, Viral
  • Genome, Viral
  • Plasmids
  • Protein Binding
  • Transcription, Genetic
  • Viral Proteins
  • Virus Replication
  • Journal Article
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

Fingerprint

Dive into the research topics of 'Genome wide, supercoiling-dependent in vivo binding of a viral protein involved in DNA replication and transcriptional control'. Together they form a unique fingerprint.

Cite this