Nucleic Acids Research, Vol 27, Issue 3 778-787, Copyright © 1999 by Oxford University Press
SS Leung and DJ Koslowsky
All guide RNAs (gRNAs) identified to date have defined 5' anchor sequences,
guiding sequences and a non-encoded 3' uridylate tail. The 5' anchor is
required for in vitro editing and is thought to be responsible for
selection and binding to the pre-edited mRNA. Little is known, however,
about how the gRNAs are used to direct RNA editing. Utilizing the
photo-reactive crosslinking agent, azidophenacyl (APA), attached to the 5'-
or 3'-terminus of the gRNA, we have begun to map the structural
relationships between the different defined regions of the gRNA with the
pre-edited mRNA. Analyses of crosslinked conjugates produced with a
5'-terminal APA group confirm that the anchor of the gRNA is correctly
positioning the interacting molecules. 3' Crosslinks (X-linker placed at
the 3'-end of a U10tail) have also been mapped for three different
gRNA/mRNA pairs. In all cases, analyses indicate that the U-tail can
interact with a range of nucleotides located upstream of the first edited
site. It appears that the U-tail prefers purine-rich sites, close to the
first few editing sites. These results suggest that the U-tail may act in
concert with the anchor to melt out secondary structure in the mRNA in the
immediate editing domain, possibly increasing the accessibility of the
editing complex to the proper editing sites.
ARTICLES
Mapping contacts between gRNA and mRNA in trypanosome RNA editing
Department of Microbiology, Michigan State University, East Lansing, MI 48824, USA.
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