Cotranscriptional recruitment of the pseudouridylsynthetase Cbf5p and of the RNA binding protein Naf1p during H/ACA snoRNP assembly.
H/ACA small nucleolar ribonucleoprotein particles (snoRNPs) are essential for the maturation and pseudouridylation of the precursor of rRNAs and other stable RNAs. Although the RNA and protein components of these RNPs have been identified, the mechanisms by which they are assembled in vivo are poorly understood. Here we show that the RNA binding protein Naf1p, which is required for H/ACA snoRNPs stability, associates with RNA polymerase II-associated proteins Spt16p, Tfg1p, and Sub1p and with H/ACA snoRNP proteins. Chromatin immunoprecipitation experiments show that Naf1p and the pseudouridylsynthetase Cbf5p cross-link specifically with the chromatin of H/ACA small nucleolar RNA (snoRNA) genes. Naf1p and Cbf5p cross-link predominantly with the 3' end of these genes, in a pattern similar to that observed for transcription elongation factor Spt16p. Cross-linking of Naf1p to H/ACA snoRNA genes requires active transcription and intact H/ACA snoRNA sequences but does not require the RNA polymerase II CTD kinase Ctk1p. These results suggest that Naf1p and Cbf5p are recruited in a cotranscriptional manner during H/ACA snoRNP assembly, possibly by binding to the nascent H/ACA snoRNA transcript during elongation or termination of transcription of H/ACA snoRNA genes.
Yang
Pok Kwan
PK
Department of Chemistry and Biochemistry and Molecular Biology Institute, UCLA, Box 951569, Los Angeles, CA 90095-1569, USA.
Hoareau
Coralie
C
Froment
Carine
C
Monsarrat
Bernard
B
Henry
Yves
Y
Chanfreau
Guillaume
G
D018797:Cell Cycle Proteins / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D004268:DNA-Binding Proteins / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D012319:RNA Polymerase II / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000235:genetics / Q000378:metabolism*; D020718:Ribonucleoproteins, Small Nucleolar / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics / Q000502:physiology*; D035602:Transcriptional Elongation Factors
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; D017411:Ribonucleoproteins, Small Nuclear; D020718:Ribonucleoproteins, Small Nucleolar; C071238:SPT16 protein, S cerevisiae; C101206:SUB1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D012319:RNA Polymerase II; D006836:Hydro-Lyases; C082278:CBF5 protein, S cerevisiae