Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein

Abstract Archaeal RNA:pseudouridine-synthase (PUS) Cbf5 in complex with proteins L7Ae, Nop10 and Gar1, and guide box H/ACA sRNAs forms ribonucleoprotein (RNP) catalysts that insure the conversion of uridines into pseudouridines (Ψs) in ribosomal RNAs (rRNAs). Nonetheless, in the absence of guide RNA...

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Autores principales:	Ryosuke Fujikane, Isabelle Behm-Ansmant, Anne-Sophie Tillault, Christine Loegler, Valérie Igel-Bourguignon, Evelyne Marguet, Patrick Forterre, Christiane Branlant, Yuri Motorin, Bruno Charpentier
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Kodakarensis Protein L7Ae Protein Nopp140 Abyssal Pseudouridylation Medicine R Science Q
Acceso en línea:	https://doaj.org/article/8f4e286c4b614fb4bbbba9971d936c88
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spelling	oai:doaj.org-article:8f4e286c4b614fb4bbbba9971d936c882021-12-02T15:07:47ZContribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein10.1038/s41598-018-32164-02045-2322https://doaj.org/article/8f4e286c4b614fb4bbbba9971d936c882018-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-32164-0https://doaj.org/toc/2045-2322Abstract Archaeal RNA:pseudouridine-synthase (PUS) Cbf5 in complex with proteins L7Ae, Nop10 and Gar1, and guide box H/ACA sRNAs forms ribonucleoprotein (RNP) catalysts that insure the conversion of uridines into pseudouridines (Ψs) in ribosomal RNAs (rRNAs). Nonetheless, in the absence of guide RNA, Cbf5 catalyzes the in vitro formation of Ψ2603 in Pyrococcus abyssi 23S rRNA and of Ψ55 in tRNAs. Using gene-disrupted strains of the hyperthermophilic archaeon Thermococcus kodakarensis, we studied the in vivo contribution of proteins Nop10 and Gar1 to the dual RNA guide-dependent and RNA-independent activities of Cbf5 on 23S rRNA. The single-null mutants of the cbf5, nop10, and gar1 genes are viable, but display a thermosensitive slow growth phenotype. We also generated a single-null mutant of the gene encoding Pus10, which has redundant activity with Cbf5 for in vitro formation of Ψ55 in tRNA. Analysis of the presence of Ψs within the rRNA peptidyl transferase center (PTC) of the mutants demonstrated that Cbf5 but not Pus10 is required for rRNA modification. Our data reveal that, in contrast to Nop10, Gar1 is crucial for in vivo and in vitro RNA guide-independent formation of Ψ2607 (Ψ2603 in P. abyssi) by Cbf5. Furthermore, our data indicate that pseudouridylation at orphan position 2589 (2585 in P. abyssi), for which no PUS or guide sRNA has been identified so far, relies on RNA- and Gar1-dependent activity of Cbf5.Ryosuke FujikaneIsabelle Behm-AnsmantAnne-Sophie TillaultChristine LoeglerValérie Igel-BourguignonEvelyne MarguetPatrick ForterreChristiane BranlantYuri MotorinBruno CharpentierNature PortfolioarticleKodakarensisProtein L7AeProtein Nopp140AbyssalPseudouridylationMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-13 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Kodakarensis Protein L7Ae Protein Nopp140 Abyssal Pseudouridylation Medicine R Science Q
spellingShingle	Kodakarensis Protein L7Ae Protein Nopp140 Abyssal Pseudouridylation Medicine R Science Q Ryosuke Fujikane Isabelle Behm-Ansmant Anne-Sophie Tillault Christine Loegler Valérie Igel-Bourguignon Evelyne Marguet Patrick Forterre Christiane Branlant Yuri Motorin Bruno Charpentier Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
description	Abstract Archaeal RNA:pseudouridine-synthase (PUS) Cbf5 in complex with proteins L7Ae, Nop10 and Gar1, and guide box H/ACA sRNAs forms ribonucleoprotein (RNP) catalysts that insure the conversion of uridines into pseudouridines (Ψs) in ribosomal RNAs (rRNAs). Nonetheless, in the absence of guide RNA, Cbf5 catalyzes the in vitro formation of Ψ2603 in Pyrococcus abyssi 23S rRNA and of Ψ55 in tRNAs. Using gene-disrupted strains of the hyperthermophilic archaeon Thermococcus kodakarensis, we studied the in vivo contribution of proteins Nop10 and Gar1 to the dual RNA guide-dependent and RNA-independent activities of Cbf5 on 23S rRNA. The single-null mutants of the cbf5, nop10, and gar1 genes are viable, but display a thermosensitive slow growth phenotype. We also generated a single-null mutant of the gene encoding Pus10, which has redundant activity with Cbf5 for in vitro formation of Ψ55 in tRNA. Analysis of the presence of Ψs within the rRNA peptidyl transferase center (PTC) of the mutants demonstrated that Cbf5 but not Pus10 is required for rRNA modification. Our data reveal that, in contrast to Nop10, Gar1 is crucial for in vivo and in vitro RNA guide-independent formation of Ψ2607 (Ψ2603 in P. abyssi) by Cbf5. Furthermore, our data indicate that pseudouridylation at orphan position 2589 (2585 in P. abyssi), for which no PUS or guide sRNA has been identified so far, relies on RNA- and Gar1-dependent activity of Cbf5.
format	article
author	Ryosuke Fujikane Isabelle Behm-Ansmant Anne-Sophie Tillault Christine Loegler Valérie Igel-Bourguignon Evelyne Marguet Patrick Forterre Christiane Branlant Yuri Motorin Bruno Charpentier
author_facet	Ryosuke Fujikane Isabelle Behm-Ansmant Anne-Sophie Tillault Christine Loegler Valérie Igel-Bourguignon Evelyne Marguet Patrick Forterre Christiane Branlant Yuri Motorin Bruno Charpentier
author_sort	Ryosuke Fujikane
title	Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
title_short	Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
title_full	Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
title_fullStr	Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
title_full_unstemmed	Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein
title_sort	contribution of protein gar1 to the rna-guided and rna-independent rrna:ψ-synthase activities of the archaeal cbf5 protein
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/8f4e286c4b614fb4bbbba9971d936c88
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Contribution of protein Gar1 to the RNA-guided and RNA-independent rRNA:Ψ-synthase activities of the archaeal Cbf5 protein

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