The DNA/RNA-dependent RNA polymerase QDE-1 generates aberrant RNA and dsRNA for RNAi in a process requiring replication protein a and a DNA helicase

Yi Liu, Heng Chi Lee, Antti P. Aalto, Qiuying Yang, Shwu Shin Chang, Guocun Huang, Daniel Fisher, Joonseok Cha, Minna M. Poranen, Dennis H. Bamford

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

The production of aberrant RNA (aRNA) is the initial step in several RNAi pathways. How aRNA is produced and specifically recognized by RNA-dependent RNA polymerases (RdRPs) to generate double-stranded RNA (dsRNA) is not clear. We previously showed that in the filamentous fungus Neurospora, the RdRP QDE-1 is required for rDNA-specific aRNA production, suggesting that QDE-1 may be important in aRNA synthesis. Here we show that a recombinant QDE-1 is both an RdRP and a DNA-dependent RNA polymerase (DdRP). Its DdRP activity is much more robust than the RdRP activity and occurs on ssDNA but not dsDNA templates. We further show that Replication Protein A (RPA), a single-stranded DNA-binding complex that interacts with QDE-1, is essential for aRNA production and gene silencing. In vitro reconstitution assays demonstrate that QDE-1 can produce dsRNA from ssDNA, a process that is strongly promoted by RPA. Furthermore, the interaction between QDE-1 and RPA requires the RecQ DNA helicase QDE-3, a homolog of the human Werner/Bloom Syndrome proteins. Together, these results suggest a novel small RNA biogenesis pathway in Neurospora and a new mechanism for the production of aRNA and dsRNA in RNAi pathways.© 2010 Lee et al.

Original languageEnglish (US)
Article numbere1000496
JournalPLoS Biology
Volume8
Issue number10
DOIs
StatePublished - Oct 2010

Fingerprint

RNA-directed RNA polymerase
DNA helicases
DNA Helicases
RNA Replicase
Double-Stranded RNA
double-stranded RNA
RNA Interference
RNA
Replication Protein A
DNA
single-stranded DNA
Neurospora
Proteins
proteins
DNA-Directed RNA Polymerases
DNA-directed RNA polymerase
RecQ Helicases
Werner Syndrome
Single-Stranded DNA
Gene Silencing

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)

Cite this

The DNA/RNA-dependent RNA polymerase QDE-1 generates aberrant RNA and dsRNA for RNAi in a process requiring replication protein a and a DNA helicase. / Liu, Yi; Lee, Heng Chi; Aalto, Antti P.; Yang, Qiuying; Chang, Shwu Shin; Huang, Guocun; Fisher, Daniel; Cha, Joonseok; Poranen, Minna M.; Bamford, Dennis H.

In: PLoS Biology, Vol. 8, No. 10, e1000496, 10.2010.

Research output: Contribution to journalArticle

Liu, Yi ; Lee, Heng Chi ; Aalto, Antti P. ; Yang, Qiuying ; Chang, Shwu Shin ; Huang, Guocun ; Fisher, Daniel ; Cha, Joonseok ; Poranen, Minna M. ; Bamford, Dennis H. / The DNA/RNA-dependent RNA polymerase QDE-1 generates aberrant RNA and dsRNA for RNAi in a process requiring replication protein a and a DNA helicase. In: PLoS Biology. 2010 ; Vol. 8, No. 10.
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