Vaccinia virus D10 protein has mRNA decapping activity, providing a mechanism for control of host and viral gene expression.
Previous studies indicated that the vaccinia virus D10 protein, which is conserved in all sequenced poxviruses, participates in the rapid turnover of host and viral mRNAs. D10 contains a motif present in the family of Nudix/MutT enzymes, a subset of which has been shown to enhance mRNA turnover in eukaryotic cells through cleavage of the 5' cap (m7GpppNm-). Here, we demonstrate that a purified recombinant D10 fusion protein possesses an intrinsic activity that liberates m7GDP from capped RNA substrates. Furthermore, point mutations in the Nudix/MutT motif abolished decapping activity. D10 has a strong affinity for capped RNA substrates (Km approximately 3 nm). RNAs of 24-309 nt were decapped to comparable extents, whereas the cap of a 12-nt RNA was uncleaved. At large molar ratios relative to capped RNA substrate, competitor m7GpppG, m7GTP, or m7GDP inhibited decapping, whereas even higher concentrations of unmethylated analogs did not. High concentrations of uncapped RNA were also inhibitory, suggesting that D10 recognizes its substrate through interaction with both cap and RNA moieties. Thus far, poxviruses represent the only virus family shown to encode a Nudix hydrolase-decapping enzyme. Although it may seem self-destructive for a virus to encode a decapping and a capping enzyme, accelerated mRNA turnover helps eliminate competing host mRNAs and allows stage-specific synthesis of viral proteins.
Parrish
Susan
S
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892-3210, USA.
Resch
Wolfgang
W
Moss
Bernard
B
D001665:Binding Sites; D005786:Gene Expression Regulation; D015967:Gene Expression Regulation, Viral; D009708:Nucleotidases / Q000502:physiology*; D017354:Point Mutation; D011755:Pyrophosphatases / Q000235:genetics; D012314:RNA Cap Analogs / Q000378:metabolism / Q000494:pharmacology; D012315:RNA Caps / Q000378:metabolism*; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D013379:Substrate Specificity; D014616:Vaccinia virus / Q000737:chemistry*; D014764:Viral Proteins / Q000378:metabolism* / Q000502:physiology; D000096964:Nudix Hydrolases
D009708:Nucleotidases; D011755:Pyrophosphatases; D012314:RNA Cap Analogs; D012315:RNA Caps; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D000096964:Nudix Hydrolases; C017516:7-methylguanosine triphosphate; C519568:D10 protein, vaccinia virus