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Functional analyses of RNA helicases in human ribosome biogenesis

dc.contributor.advisorBohnsack, Markus Prof. Dr.
dc.contributor.authorChoudhury, Priyanka
dc.titleFunctional analyses of RNA helicases in human ribosome biogenesisde
dc.contributor.refereeBohnsack, Markus Prof. Dr.
dc.description.abstractengEukaryotic ribosomes are essential ribonucleoprotein (RNP) complexes consisting of four ribosomal RNAs (18S, 5S, 5.8S, 28S/25S) and approximately 80 ribosomal proteins, which are required for the process of translating the information encoded in mRNAs into proteins. Ribosome biogenesis requires a dynamic interplay between a multitude of trans-acting biogenesis factors such as ATPases, GTPases, snoRNAs, nucleases and RNA helicases, which assist in the maturation of pre-ribosomal subunits. RNA helicases are RNA-dependent NTPases that play key roles in all aspects of RNA metabolism including pre-mRNA splicing, translation, transcription and ribosome biogenesis. The majority of RNA helicases involved in ribosome biogenesis belong to either the DEAD- or DEAH-box family and consist of a structurally conserved helicase core flanked by N- and C-terminal domains that can serve as platforms for protein-protein/protein-RNA interaction. In yeast, 21 RNA helicases are implicated in the process of ribosome biogenesis and likely play key roles in the structural remodelling of pre-ribosomal subunits. Genome-wide screens and proteomic analyses of mammalian nucleoli have suggested the involvement of many orthologues of yeast RNA helicases in the human ribosome assembly pathway as well as highlighting additional helicases that may regulate the process in humans. Although several RNA helicases are implicated in human ribosome assembly, the precise physiological roles and in vivo substrates of these helicases are often not well-characterized. In this study, we aimed to gain insights into the roles of two RNA helicases-DHX37 and DDX55, which are implicated in SSU and LSU biogenesis respectively. An RNAi-based rescue system indicated that the catalytic activity of DHX37 is required for the effective maturation of 18S rRNA. Using an in vivo crosslinking approach, we show that DHX37 crosslinks to the U3 snoRNA and that expression of a catalytically inactive mutant of DHX37 leads to the accumulation the pre-ribosomal accumulation of U3, suggesting a role for DHX37 in release of the U3 snoRNP from pre-ribosomal particles. In addition, we identified UTP14A as a cofactor of DHX37, and our data suggest that UTP14A and DHX37 act in a synergistic manner to promote the dissociation of U3 snoRNA from pre-ribosomal subunits. While our data implied that DHX37 mediated release of U3 snoRNP is conserved in eukaryotes, we also revealed several interesting differences between the yeast and human proteins in terms of recruitment and function of the helicase during late steps of ribosome assembly. Furthermore, our crosslinking data also identified a binding site of DDX55 on the 28S rRNA sequence, consistent with its requirement for LSU maturation. DDX55 was found to predominantly localize in the nucleoplasm suggesting a role in late-stage ribosome biogenesis. We further investigated the function of the lysine-rich C-terminal tail of DDX55 and our results indicate a possible role of the C-terminal tail in the recruitment of DDX55 to pre-ribosomal complexes. Taken together, our results provide important new insights into the roles of RNA helicases in human ribosome
dc.contributor.coRefereeRodnina, Marina Prof. Dr.
dc.contributor.thirdRefereeStülke, Jörg Prof. Dr.
dc.contributor.thirdRefereeFicner, Ralf Prof. Dr.
dc.contributor.thirdRefereeMeinecke, Michael Prof. Dr.
dc.contributor.thirdRefereeStein, Alexander Dr.
dc.subject.engRibosome Biogenesisde
dc.subject.engRNA helicasesde
dc.affiliation.instituteGöttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB)de
dc.subject.gokfullBiologie (PPN619462639)de

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