Coupled GTPase and remodelling ATPase activities form a checkpoint for ribosome export.
Eukaryotic ribosomes are assembled by a complex pathway that extends from the nucleolus to the cytoplasm and is powered by many energy-consuming enzymes. Nuclear export is a key, irreversible step in pre-ribosome maturation, but mechanisms underlying the timely acquisition of export competence remain poorly understood. Here we show that a conserved Saccharomyces cerevisiae GTPase Nug2 (also known as Nog2, and as NGP-1, GNL2 or nucleostemin 2 in human) has a key role in the timing of export competence. Nug2 binds the inter-subunit face of maturing, nucleoplasmic pre-60S particles, and the location clashes with the position of Nmd3, a key pre-60S export adaptor. Nug2 and Nmd3 are not present on the same pre-60S particles, with Nug2 binding before Nmd3. Depletion of Nug2 causes premature Nmd3 binding to the pre-60S particles, whereas mutations in the G-domain of Nug2 block Nmd3 recruitment, resulting in severe 60S export defects. Two pre-60S remodelling factors, the Rea1 ATPase and its co-substrate Rsa4, are present on Nug2-associated particles, and both show synthetic lethal interactions with nug2 mutants. Release of Nug2 from pre-60S particles requires both its K(+)-dependent GTPase activity and the remodelling ATPase activity of Rea1. We conclude that Nug2 is a regulatory GTPase that monitors pre-60S maturation, with release from its placeholder site linked to recruitment of the nuclear export machinery.
Biochemie-Zentrum der Universität Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany.
Yoshitaka
Y
Matsuo
Wellcome Trust Centre for Cell Biology, The University of Edinburgh, Edinburgh UK.|Centre for Synthetic and Systems Biology (SynthSys), University of Edinburgh, Edinburgh, UK.
Sander
S
Granneman
Biochemie-Zentrum der Universität Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany.
Matthias
M
Thoms
Biochemie-Zentrum der Universität Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany.
Rizos-Georgios
RG
Manikas
Wellcome Trust Centre for Cell Biology, The University of Edinburgh, Edinburgh UK.
David
D
Tollervey
Biochemie-Zentrum der Universität Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany.
Ed
E
Hurt
D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005804:Genes, Lethal / Q000235:genetics; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D011188:Potassium / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000235:genetics; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D012269:Ribosomal Proteins / Q000378:metabolism; D054683:Ribosome Subunits, Large, Eukaryotic / Q000737:chemistry / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C117894:NMD3 protein, S cerevisiae; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; C504806:Rsa4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000251:Adenosine Triphosphatases; D020558:GTP Phosphohydrolases; C477781:NOG2 protein, S cerevisiae; C496538:MDN1 protein, S cerevisiae; D000074183:ATPases Associated with Diverse Cellular Activities; D011188:Potassium