Abstract
Box C/D small nucleolar RNAs (snoRNAs) direct site-specific methylation of ribose 2'-hydroxyls in ribosomal and spliceosomal RNAs. To identify snoRNA functional groups contributing to assembly of an active box C/D snoRNP in Xenopus oocytes, we developed an in vivo nucleotide analog interference mapping procedure. Deleterious substitutions consistent with requirements for binding the 15.5 kD protein clustered within the terminal box C/D motif only. In vitro analyses confirmed a single interaction site for recombinant 15.5 kD protein and identified the exocyclic amine of A89 in box D as essential for binding. Our results argue that the 15.5 kD protein interacts asymmetrically with the two sets of conserved box C/D elements and that its binding is primarily responsible for the stability of box C/D snoRNAs in vivo.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Adenosine / analogs & derivatives
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Adenosine / metabolism
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Animals
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Base Sequence
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Conserved Sequence
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Methylation
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Models, Molecular
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Molecular Sequence Data
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Molecular Weight
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Nucleic Acid Conformation
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Oocytes
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Protein Binding
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RNA, Small Nucleolar / chemistry
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RNA, Small Nucleolar / genetics
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RNA, Small Nucleolar / metabolism*
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RNA-Binding Proteins / chemistry
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RNA-Binding Proteins / metabolism*
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Ribonucleoproteins, Small Nuclear / chemistry
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Ribonucleoproteins, Small Nuclear / metabolism*
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Thionucleotides / chemistry
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Thionucleotides / metabolism
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Xenopus / metabolism
Substances
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RNA, Small Nucleolar
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RNA-Binding Proteins
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Recombinant Proteins
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Ribonucleoproteins, Small Nuclear
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Thionucleotides
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Adenosine