Rrp5 binding at multiple sites coordinates pre-rRNA processing and assembly.
In vivo UV crosslinking identified numerous preribosomal RNA (pre-rRNA) binding sites for the large, highly conserved ribosome synthesis factor Rrp5. Intramolecular complementation has shown that the C-terminal domain (CTD) of Rrp5 is required for pre-rRNA cleavage at sites A0-A2 on the pathway of 18S rRNA synthesis, whereas the N-terminal domain (NTD) is required for A3 cleavage on the pathway of 5.8S/25S rRNA synthesis. The CTD was crosslinked to sequences flanking A2 and to the snoRNAs U3, U14, snR30, and snR10, which are required for cleavage at A0-A2. The NTD was crosslinked to sequences flanking A3 and to the RNA component of ribonuclease MRP, which cleaves site A3. Rrp5 could also be directly crosslinked to several large structural proteins and nucleoside triphosphatases. A key role in coordinating preribosomal assembly and processing was confirmed by chromatin spreads. Following depletion of Rrp5, cotranscriptional cleavage was lost and preribosome compaction greatly reduced.
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland.
Simon
S
Lebaron
Department of Molecular Biosciences, WGI, Stockholm University, 106 91 Stockholm, Sweden.
Asa
A
Segerstolpe
Department of Microbiology, Immunology and Cancer Biology, University of Virginia Health System, Charlottesville, VA 22908-0734, USA.
Sarah L
SL
French
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland.
Tatiana
T
Dudnakova
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland.
Flavia
F
de Lima Alves
SynthSys, JR Waddington Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland.
Sander
S
Granneman
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland.
Juri
J
Rappsilber
Department of Microbiology, Immunology and Cancer Biology, University of Virginia Health System, Charlottesville, VA 22908-0734, USA.
Ann L
AL
Beyer
Department of Molecular Biosciences, WGI, Stockholm University, 106 91 Stockholm, Sweden.
Lars
L
Wieslander
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, Kings Buildings, Mayfield Road, Edinburgh EH9 3JR, Scotland. Electronic address: d.tollervey@ed.ac.uk.
David
D
Tollervey
D001483:Base Sequence; D001665:Binding Sites; D004722:Endoribonucleases / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D043583:Nucleoside-Triphosphatase / Q000235:genetics / Q000378:metabolism; D012322:RNA Precursors / Q000235:genetics* / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012331:RNA, Fungal / Q000235:genetics*; D012335:RNA, Ribosomal / Q000235:genetics* / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics*; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
D005656:Fungal Proteins; D012322:RNA Precursors; D012331:RNA, Fungal; D012335:RNA, Ribosomal; D004722:Endoribonucleases; C058330:mitochondrial RNA-processing endoribonuclease; D043583:Nucleoside-Triphosphatase