title: Release of U18 snoRNA from its host intron requires interaction of Nop1p with the Rnt1p endonuclease.
abstract: An external stem, essential for the release of small nucleolar RNAs (snoRNAs) from their pre-mRNAs, flanks the majority of yeast intron-encoded snoRNAs. Even if this stem is not a canonical Rnt1p substrate, several experiments have indicated that the Rnt1p endonuclease is required for snoRNA processing. To identify the factors necessary for processing of intron-encoded snoRNAs, we have raised in vitro extracts able to reproduce such activity. We found that snoRNP factors are associated with the snoRNA- coding region throughout all the processing steps, and that mutants unable to assemble snoRNPs have a processing-deficient phenotype. Specific depletion of Nop1p completely prevents U18 snoRNA synthesis, but does not affect processing of a dicistronic snoRNA-coding unit that has a canonical Rnt1p site. Correct cleavage of intron-encoded U18 and snR38 snoRNAs can be reproduced in vitro by incubating together purified Nop1p and Rnt1p. Pull-down experiments showed that the two proteins interact physically. These data indicate that cleavage of U18, snR38 and possibly other intron-encoded snoRNAs is a regulated process, since the stem is cleaved by the Rnt1p endonuclease only when snoRNP assembly has occurred.
authors.affiliation: Institute Pasteur Fondazione Cenci-Bolognetti, Department of Genetics and Molecular Biology, University La Sapienza, P. le A. Moro 5, 00185 Rome, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Giorgi
authors.forename: A
authors.initials: A
authors.lastname: Fatica
authors.forename: R
authors.initials: R
authors.lastname: Nagel
authors.forename: I
authors.initials: I
authors.lastname: Bozzoni
mesh_terms: D017931:DNA Primers / Q000378:metabolism; D004722:Endoribonucleases / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D005796:Genes; D005982:Glutathione Transferase / Q000378:metabolism; D007438:Introns; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D010641:Phenotype; D011485:Protein Binding; D020537:RNA, Small Nucleolar / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D043244:Ribonuclease III; D020718:Ribonucleoproteins, Small Nucleolar; D029701:Saccharomyces cerevisiae Proteins
chemical_list: D017931:DNA Primers; D005656:Fungal Proteins; C064793:NOP1 protein, S cerevisiae; D009687:Nuclear Proteins; D020537:RNA, Small Nucleolar; D011993:Recombinant Fusion Proteins; D020718:Ribonucleoproteins, Small Nucleolar; D029701:Saccharomyces cerevisiae Proteins; D005982:Glutathione Transferase; D004722:Endoribonucleases; D043244:Ribonuclease III