title: A conserved major groove antideterminant for Saccharomyces cerevisiae RNase III recognition.
abstract: Rnt1p, the only known Saccharomyces cerevisiae RNase III double-stranded RNA endonuclease, plays important roles in the processing of precursors of ribosomal RNAs and small nuclear and nucleolar RNAs and in the surveillance of unspliced pre-mRNAs. Specificity of cleavage by Rnt1p relies on the presence of RNA tetraloop structures with the consensus sequence AGNN at the top of the target dsRNA. The sequences of 79 fungal RNase III substrates were inspected to identify additional conserved sequence elements or antideterminants that may contribute to Rnt1p recognition of the double-stranded RNA. Surprisingly, U-A sequences at the base pair adjacent to the conserved terminal tetraloop (closing base pair) were found to be absent from all but one inspected sequence. Analysis of chemically modified variants of the closing base pair showed that the presence of exocyclic groups in the major groove of purines 3' to the last nucleotide of the tetraloop inhibits Rnt1p cleavage without strongly inhibiting Rnt1p binding. We propose that these groups interfere with the recognition of the RNA substrate by the catalytic domain of Rnt1p. These results identify exocyclic groups of purines in the major groove downstream of the tetraloop as a major antideterminant in S. cerevisiae RNase III activity, and suggest a rationale for their apparent counter selection in RNA processing sites.
authors.affiliation: Department of Chemistry and Biochemistry and Molecular Biology Institute, Box 951569, University of California, Los Angeles, California 90095-1569, USA.
authors.forename: Mui
authors.initials: M
authors.lastname: Sam
authors.forename: Anthony K
authors.initials: AK
authors.lastname: Henras
authors.forename: Guillaume
authors.initials: G
authors.lastname: Chanfreau
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000378:metabolism; D020029:Base Pairing / Q000235:genetics; D001483:Base Sequence; D020134:Catalytic Domain / Q000235:genetics; D017124:Conserved Sequence; D004249:DNA Damage; D004791:Enzyme Inhibitors / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics; D012331:RNA, Fungal / Q000737:chemistry* / Q000378:metabolism*; D043244:Ribonuclease III / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D014529:Uridine / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; C468367:RNT1 protein, S cerevisiae; D043244:Ribonuclease III; D000225:Adenine; D014529:Uridine