RNA 3'-end mismatch excision by the severe acute respiratory syndrome coronavirus nonstructural protein nsp10/nsp14 exoribonuclease complex

Proc Natl Acad Sci U S A. 2012 Jun 12;109(24):9372-7. doi: 10.1073/pnas.1201130109. Epub 2012 May 25.

Abstract

The replication/transcription complex of severe acute respiratory syndrome coronavirus is composed of at least 16 nonstructural proteins (nsp1-16) encoded by the ORF-1a/1b. This complex includes replication enzymes commonly found in positive-strand RNA viruses, but also a set of RNA-processing activities unique to some nidoviruses. The nsp14 protein carries both exoribonuclease (ExoN) and (guanine-N7)-methyltransferase (N7-MTase) activities. The nsp14 ExoN activity ensures a yet-uncharacterized function in the virus life cycle and must be regulated to avoid nonspecific RNA degradation. In this work, we show that the association of nsp10 with nsp14 stimulates >35-fold the ExoN activity of the latter while playing no effect on N7-MTase activity. Nsp10 mutants unable to interact with nsp14 are not proficient for ExoN activation. The nsp10/nsp14 complex hydrolyzes double-stranded RNA in a 3' to 5' direction as well as a single mismatched nucleotide at the 3'-end mimicking an erroneous replication product. In contrast, di-, tri-, and longer unpaired ribonucleotide stretches, as well as 3'-modified RNAs, resist nsp10/nsp14-mediated excision. In addition to the activation of nsp16-mediated 2'-O-MTase activity, nsp10 also activates nsp14 in an RNA processing function potentially connected to a replicative mismatch repair mechanism.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Pair Mismatch*
  • Exoribonucleases / genetics
  • Exoribonucleases / metabolism*
  • Open Reading Frames
  • RNA Processing, Post-Transcriptional
  • RNA, Viral / genetics*
  • Severe acute respiratory syndrome-related coronavirus / genetics*
  • Viral Nonstructural Proteins / genetics
  • Viral Nonstructural Proteins / metabolism*

Substances

  • Nsp10 protein, SARS virus
  • RNA, Viral
  • Viral Nonstructural Proteins
  • nsp14 protein, SARS coronavirus
  • Exoribonucleases