title: DEAD-box RNA helicase Dbp4 is required for small-subunit processome formation and function.
abstract: DEAD-box RNA helicase Dbp4 is required for 18S rRNA synthesis: cellular depletion of Dbp4 impairs the early cleavage reactions of the pre-rRNA and causes U14 small nucleolar RNA (snoRNA) to remain associated with pre-rRNA. Immunoprecipitation experiments (IPs) carried out with whole-cell extracts (WCEs) revealed that hemagglutinin (HA)-tagged Dbp4 is associated with U3 snoRNA but not with U14 snoRNA. IPs with WCEs also showed association with the U3-specific protein Mpp10, which suggests that Dbp4 interacts with the functionally active U3 RNP; this particle, called the small-subunit (SSU) processome, can be observed at the 5' end of nascent pre-rRNA. Electron microscopy analyses indicated that depletion of Dbp4 compromised SSU processome formation and cotranscriptional cleavage of the pre-rRNA. Sucrose density gradient analyses revealed that depletion of U3 snoRNA or the Mpp10 protein inhibited the release of U14 snoRNA from pre-rRNA, just as was seen with Dbp4-depleted cells, indicating that alteration of SSU processome components has significant consequences for U14 snoRNA dynamics. We also found that the C-terminal extension flanking the catalytic core of Dbp4 plays an important role in the release of U14 snoRNA from pre-rRNA.
authors.affiliation: Département des sciences biologiques and Centre de recherche BioMed, Université du Québec à Montréal, Montreal, Quebec, Canada.
authors.forename: Sahar
authors.initials: S
authors.lastname: Soltanieh
authors.affiliation: Department of Microbiology, Immunology and Cancer Biology, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Yvonne N
authors.initials: YN
authors.lastname: Osheim
authors.affiliation: Département des sciences biologiques and Centre de recherche BioMed, Université du Québec à Montréal, Montreal, Quebec, Canada.
authors.forename: Krasimir
authors.initials: K
authors.lastname: Spasov
authors.affiliation: Département des sciences biologiques and Centre de recherche BioMed, Université du Québec à Montréal, Montreal, Quebec, Canada.
authors.forename: Christian
authors.initials: C
authors.lastname: Trahan
authors.affiliation: Department of Microbiology, Immunology and Cancer Biology, University of Virginia Health System, Charlottesville, Virginia, USA.
authors.forename: Ann L
authors.initials: AL
authors.lastname: Beyer
authors.affiliation: Département des sciences biologiques and Centre de recherche BioMed, Université du Québec à Montréal, Montreal, Quebec, Canada dragon.francois@uqam.ca.
authors.forename: François
authors.initials: F
authors.lastname: Dragon
mesh_terms: D002499:Centrifugation, Density Gradient; D002843:Chromatin / Q000737:chemistry; D053487:DEAD-box RNA Helicases / Q000378:metabolism*; D005838:Genotype; D008854:Microscopy, Electron; D010750:Phosphoproteins / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D020365:RNA Helicases / Q000378:metabolism; D012316:RNA Nucleotidyltransferases / Q000378:metabolism*; D012337:RNA, Ribosomal, 18S / Q000378:metabolism; D020537:RNA, Small Nucleolar / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D012270:Ribosomes / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
chemical_list: D002843:Chromatin; C111098:MPP10 protein, S cerevisiae; D010750:Phosphoproteins; D012337:RNA, Ribosomal, 18S; D020537:RNA, Small Nucleolar; C117378:RNA, U3 small nucleolar; D012261:Ribonucleoproteins; D029701:Saccharomyces cerevisiae Proteins; D012316:RNA Nucleotidyltransferases; C106767:HCA4 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases