Article

Prp43 Bound at Different Sites on the Pre-rRNA Performs Distinct Functions in Ribosome Synthesis

Wellcome Trust Centre for Cell Biology, University of Edinburgh, UK.
Molecular cell (Impact Factor: 14.46). 11/2009; 36(4):583-92. DOI: 10.1016/j.molcel.2009.09.039
Source: PubMed

ABSTRACT Yeast ribosome synthesis requires 19 different RNA helicases, but none of their pre-rRNA-binding sites were previously known, making their precise functions difficult to determine. Here we identify multiple binding sites for the helicase Prp43 in the 18S and 25S rRNA regions of pre-rRNAs, using UV crosslinking. Binding in 18S was predominantly within helix 44, close to the site of 18S 3' cleavage, in which Prp43 is functionally implicated. Four major binding sites were identified in 25S, including helix 34. In strains depleted of Prp43 or expressing only catalytic point mutants, six snoRNAs that guide modifications close to helix 34 accumulated on preribosomes, implicating Prp43 in their release, whereas other snoRNAs showed reduced preribosome association. Prp43 was crosslinked to snoRNAs that target sequences close to its binding sites, indicating direct interactions. We propose that Prp43 acts on preribosomal regions surrounding each binding site, with distinct functions at different locations.

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    • "was performed as previously described (Bohnsack et al. 2009; Granneman et al. 2009; Bohnsack et al. 2012). In brief, UV crosslinking was performed in living cells either in culture medium (in culturo) or pelleted and resuspended in small volume (in vivo), and Rok1-containing complexes were isolated first on IgG sepharose, eluted with TEV protease, and purified on Nickel-NTA. "
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    • "The unexpected detection of C2ORF3 protein also in the nucleoli suggests a function for C2ORF3 in rRNA processing . As the hPrp43 protein is shown to be involved in rRNA processing (Bohnsack et al. 2009; Lebaron et al. 2009; Pertschy et al. 2009), it is quite conceivable that the C2ORF3 protein would function in this process in collaboration with hPrp43. It would be interesting to determine whether the knockdown of C2ORF3 protein affects ribosomal RNA processing and/or nucleoli structure. "
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