title: The PIN domain endonuclease Utp24 cleaves pre-ribosomal RNA at two coupled sites in yeast and humans.
abstract: During ribosomal RNA (rRNA) maturation, cleavages at defined sites separate the mature rRNAs from spacer regions, but the identities of several enzymes required for 18S rRNA release remain unknown. PilT N-terminus (PIN) domain proteins are frequently endonucleases and the PIN domain protein Utp24 is essential for early cleavages at three pre-rRNA sites in yeast (A0, A1 and A2) and humans (A0, 1 and 2a). In yeast, A1 is cleaved prior to A2 and both cleavages require base-pairing by the U3 snoRNA to the central pseudoknot elements of the 18S rRNA. We found that yeast Utp24 UV-crosslinked in vivo to U3 and the pseudoknot, placing Utp24 close to cleavage at site A1. Yeast and human Utp24 proteins exhibited in vitro endonuclease activity on an RNA substrate containing yeast site A2. Moreover, an intact PIN domain in human UTP24 was required for accurate cleavages at sites 1 and 2a in vivo, whereas mutation of another potential site 2a endonuclease, RCL1, did not affect 18S production. We propose that Utp24 cleaves sites A1/1 and A2/2a in yeast and human cells.
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: Graeme R
authors.initials: GR
authors.lastname: Wells
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: Franziska
authors.initials: F
authors.lastname: Weichmann
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: David
authors.initials: D
authors.lastname: Colvin
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: Katherine E
authors.initials: KE
authors.lastname: Sloan
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3JR, UK MRC Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh EH4 2XU, UK.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Kudla
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3JR, UK.
authors.forename: David
authors.initials: D
authors.lastname: Tollervey
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Watkins
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4HH, UK Claudia.Schneider@ncl.ac.uk.
authors.forename: Claudia
authors.initials: C
authors.lastname: Schneider