A RAC Protein-binding Site in the Internal Transcribed
Spacer 2 of Pre-rRNA Transcripts from Schizosaccharomyces
pombe*
Priyanka D.
Abeyrathne,
Atanas I.
Lalev, and
Ross N.
Nazar
From the Department of Molecular Biology and Genetics,
University of Guelph, Guelph, Ontario N1G 2W1, Canada
The interdependence of steps in the processing of
the eukaryotic preribosomal rRNA transcripts indicate that rRNA
processing, at least in part, acts as a quality control mechanism to
help ensure that only functional rRNA is incorporated into mature
ribosomes. In search of structural components that underlie this
interdependence, we have isolated a large protein complex or RAC that
contains an independent binding site for all four of the transcribed
spacers in the nascent pre-rRNA. In this study the RAC-binding site in the internal transcribed spacer 2 sequence of Schizosaccharomyces pombe rRNA transcripts was identified, and the influence of this site on rRNA maturation was assessed. Modification exclusion analyses indicate that the protein complex interacts with a helical domain previously shown to contain features common to both the internal transcribed spacer 1 and the 3'-external transcribed spacer.
Mutagenic analyses in vitro confirm an interaction with
this sequence, and parallel analyses in vivo indicated a
critical role in both the maturation of the rRNA components of the
large subunit as well as the 18 S rRNA component of the small subunit.
Hybridization analyses also indicated greatly elevated levels of
unprocessed nascent RNA. These effects are contrasted with mutations in
other regions of the secondary structure that resulted in some
reduction of plasmid-derived mature rRNA but no elevated levels of the
precursor molecules. The significance with respect to rRNA maturation
and the interdependences in rRNA processing are discussed.
*
This work was supported by the Natural Sciences and
Engineering Research Council of Canada.The costs of publication of this article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
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