Nucleic Acids Research, Vol 27, Issue 20 3986-3994, Copyright © 1999 by Oxford University Press
M Bell and A Bindereif
We have cloned the single-copy gene for the trans -spliceosomal U5 snRNA
from the trypanosomatid species Leptomonas seymouri, using U5 RNA affinity
selection and cDNA cloning. Sequence comparison revealed that the trans
-spliceosomal U5 RNAs from trypanosomatid species share certain
characteristic features. Interestingly, the affinity selection procedure
yielded-in addition to the bona fide U5 RNA-a closely related small RNA,
which can be folded into the same secondary structure, but carries three
changes in the loop sequence. This raises the possibility that there may be
a larger family of U5-like RNAs in trypanosomes. To study the U5 snRNP
assembly and function in trypanosomes we have established a stable
expression system in L.seymouri. Two cell lines have been generated that
express U5 RNAs with mutations in the Sm site, resulting in a defect of
core snRNP formation. In addition, the U5 Sm- mutant RNAs behaved
differently in cell fractionation, implying a defect in nuclear
localization. In sum, this demonstrates for the first time that the Sm site
of trypanosome snRNAs contributes an essential element for stable core RNP
assembly and may be important for nuclear localization, in analogy to the
Sm site function of cis -spliceosomal snRNAs in higher eucaryotes.
ARTICLES
Cloning and mutational analysis of the Leptomonas seymouri U5 snRNA gene: function of the Sm site in core RNP formation and nuclear localization
Institut fur Biochemie, Humboldt-Universitat/Charite, Monbijoustrasse 2, D-10117 Berlin, Germany.
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