RNA 3'-end mismatch excision by the severe acute respiratory syndrome coronavirus nonstructural protein nsp10/nsp14 exoribonuclease complex.
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.
Architecture et Fonction des Macromolécules Biologiques, Unité Mixte de Recherche 7257, Ecole Supérieure d'Ingénieurs de Luminy Case 925,  Centre  National de la Recherche Scientifique and Aix-Marseille Université, 13288 Marseille,  France.
Mickaël
M
Bouvet
Isabelle
I
Imbert
Lorenzo
L
Subissi
Laure
L
Gluais
Bruno
B
Canard
Etienne
E
Decroly
D020137:Base Pair Mismatch; D005095:Exoribonucleases / Q000235:genetics / Q000378:metabolism*; D016366:Open Reading Frames; D012323:RNA Processing, Post-Transcriptional; D012367:RNA, Viral / Q000235:genetics*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics*; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*
C558878:Nsp10 protein, SARS virus; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; C525596:nsp14 protein, SARS coronavirus; D005095:Exoribonucleases