Facilitated recycling protects human RNA polymerase III from repression by Maf1 in vitro

Pavel Čabart, Jae Hoon Lee, Ian M. Willis

Research output: Contribution to journalArticle

36 Scopus citations

Abstract

Yeast cells synthesize ∼3-6 million molecules of tRNA every cell cycle at a rate of ∼2-4 transcripts/gene/s. This high rate of transcription is achieved through many rounds of reinitiation by RNA polymerase (pol) III on stable DNA-bound complexes of the initiation factor TFIIIB. Studies in yeast have shown that the rate of reinitiation is increased by facilitated recycling, a process that involves the repeated reloading of the polymerase on the same transcription unit. However, when nutrients become limiting or stress conditions are encountered, RNA pol III transcription is rapidly repressed through the action of the conserved Maf1 protein. Here we examine the relationship between Maf1-mediated repression and facilitated recycling in a human RNA pol III in vitro system. Using an immobilized template transcription assay, we demonstrate that facilitated recycling is conserved from yeast to humans. We assessed the ability of recombinant human Maf1 to inhibit different steps in transcription before and after preinitiation complex assembly. We show that recombinant Maf1 can inhibit the recruitment of TFIIIB and RNA pol III to immobilized templates. However, RNA pol III bound to preinitiation complexes or in elongation complexes is protected from repression by Maf1 and can undergo several rounds of initiation. This indicates that recombinant Maf1 is unable to inhibit facilitated recycling. The data suggest that additional biochemical stepsmaybe necessary for rapid Maf1-dependent repression of RNA pol III transcription.

Original languageEnglish (US)
Pages (from-to)36108-36117
Number of pages10
JournalJournal of Biological Chemistry
Volume283
Issue number52
DOIs
StatePublished - Dec 26 2008

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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