Reconstitution of human rRNA gene transcription in mouse cells by a complete SL1 complex

Kensaku Murano, Mitsuru Okuwaki, Fumitaka Momose, Michiko Kumakura, Shuhei Ueshima, Robert F. Newbold, Kyosuke Nagata

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

An important characteristic of the transcription of a ribosomal RNA gene (rDNA) mediated by DNA-dependent RNA polymerase (Pol) I is its stringent species specificity. SL1/TIF-IB is a key complex for species specificity, but its functional complex has not been reconstituted. Here, we established a novel and highly sensitive monitoring system for Pol I transcription to reconstitute the SL1 activity in which a transcript harboring a reporter gene synthesized by Pol I is amplified and converted into translatable mRNA by the influenza virus RNA-dependent RNA polymerase. Using this monitoring system, we reconstituted Pol I transcription from the human rDNA promoter in mouse cells by expressing four human TATA-binding protein (TBP)-associated factors (TAFIs) in the SL1 complex. The reconstituted SL1 also re-activated human rDNA transcription in mouse A9 cells carrying an inactive human chromosome 21 that contains the rDNA cluster. Chimeric SL1 complexes containing human and mouse TAFIs could be formed, but these complexes were inactive for human rDNA transcription. We conclude that four human TAFIs are necessary and sufficient to overcome the barrier of species specificity for human rDNA transcription in mouse cells.2014.

Original languageEnglish
Pages (from-to)3309-3319
Number of pages11
JournalJournal of Cell Science
Volume127
Issue number15
DOIs
Publication statusPublished - 2014

Keywords

  • RNA polymerase I
  • Ribosomal RNA gene
  • SL1
  • Species specificity
  • TIF-IB
  • Transcription

ASJC Scopus subject areas

  • Cell Biology

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