title: Dominant Rio1 kinase/ATPase catalytic mutant induces trapping of late pre-40S biogenesis factors in 80S-like ribosomes.
abstract: During eukaryotic ribosome biogenesis, members of the conserved atypical serine/threonine protein kinase family, the RIO kinases (Rio1, Rio2 and Rio3) function in small ribosomal subunit biogenesis. Structural analysis of Rio2 indicated a role as a conformation-sensing ATPase rather than a kinase to regulate its dynamic association with the pre-40S subunit. However, it remained elusive at which step and by which mechanism the other RIO kinase members act. Here, we have determined the crystal structure of the human Rio1-ATP-Mg(2+) complex carrying a phosphoaspartate in the active site indicative of ATPase activity. Structure-based mutations in yeast showed that Rio1's catalytic activity regulates its pre-40S association. Furthermore, we provide evidence that Rio1 associates with a very late pre-40S via its conserved C-terminal domain. Moreover, a rio1 dominant-negative mutant defective in ATP hydrolysis induced trapping of late biogenesis factors in pre-ribosomal particles, which turned out not to be pre-40S but 80S-like ribosomes. Thus, the RIO kinase fold generates a versatile ATPase enzyme, which in the case of Rio1 is activated following the Rio2 step to regulate one of the final 40S maturation events, at which time the 60S subunit is recruited for final quality control check.
authors.affiliation: Biochemistry Center, University of Heidelberg, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany Universität Regensburg, Biochemie-Zentrum Regensburg (BZR), Lehrstuhl Biochemie III, 93053 Regensburg, Germany sebastien.ferreira-cerca@ur.de.
authors.forename: Sébastien
authors.identifier: http://orcid.org/0000-0002-0522-843X
authors.initials: S
authors.lastname: Ferreira-Cerca
authors.affiliation: Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA University of Maryland Marlene and Stewart Greenebaum Cancer Center, Baltimore, MD 21201, USA.
authors.forename: Irene
authors.initials: I
authors.lastname: Kiburu
authors.affiliation: Biochemistry Center, University of Heidelberg, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany.
authors.forename: Emma
authors.initials: E
authors.lastname: Thomson
authors.affiliation: Department of Chemistry and Biochemistry, University of Maryland, College Park, MD 20742, USA University of Maryland Marlene and Stewart Greenebaum Cancer Center, Baltimore, MD 21201, USA sebastien.ferreira-cerca@ur.de.
authors.forename: Nicole
authors.initials: N
authors.lastname: LaRonde
authors.affiliation: Biochemistry Center, University of Heidelberg, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany ed.hurt@bzh.uni-heidelberg.de.
authors.forename: Ed
authors.initials: E
authors.lastname: Hurt
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D001224:Aspartic Acid / Q000737:chemistry; D020134:Catalytic Domain; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054683:Ribosome Subunits, Large, Eukaryotic / Q000378:metabolism; D054682:Ribosome Subunits, Small, Eukaryotic / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*
chemical_list: D001224:Aspartic Acid; D017346:Protein Serine-Threonine Kinases; C469385:RIOK1 protein, human; D000251:Adenosine Triphosphatases