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Molecular and Cellular Biology, August 2000, p. 5415-5424, Vol. 20, No. 15
Wellcome Trust Centre for Cell Biology, ICMB,
The University of Edinburgh, Edinburgh EH9 3JR,
Scotland,1 and GIM-Biotechnologies,
Institute Pasteur, 75724 Paris Cedex 15, France2
Received 10 March 2000/Returned for modification 10 April
2000/Accepted 2 May 2000
Almost all small eukaryotic RNAs are processed from transiently
stabilized 3'-extended forms. A key question is how and why such
intermediates are stabilized and how they can then be processed to the
mature RNA. Here we report that yeast U3 is also processed from a
3'-extended precursor. The major 3'-extended forms of U3 (U3-3'I and
-II) lack the cap trimethylation present in mature U3 and are not
associated with small nucleolar RNP (snoRNP) proteins that bind mature
U3, i.e., Nop1p, Nop56p, and Nop58p. Depletion of Nop58p leads to the
loss of mature U3 but increases the level of U3-3'I and -II, indicating
a requirement for the snoRNP proteins for final maturation. Pre-U3 is
cleaved by the endonuclease Rnt1p, but U3-3'I and -II do not extend to
the Rnt1p cleavage sites. Rather, they terminate at poly(U) tracts,
suggesting that they might be bound by Lhp1p (the yeast homologue of
La). Immunoprecipitation of Lhp1p fused to Staphylococcus
aureus protein A resulted in coprecipitation of both U3-3'I and
-II. Deletion of LHP1, which is nonessential, led to the
loss of U3-3'I and -II. We conclude that pre-U3 is cleaved by Rnt1p,
followed by exonuclease digestion to U3-3'I and -II. These species are
stabilized against continued degradation by binding of Lhp1p.
Displacement of Lhp1p by binding of the snoRNP proteins allows final
maturation, which involves the exosome complex of 3'
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Precursors to the U3 Small Nucleolar RNA Lack Small
Nucleolar RNP Proteins but Are Stabilized by La Binding

5' exonucleases.
*
Corresponding author. Mailing address: Wellcome Trust
Centre for Cell Biology, ICMB, Swann Building, King's Buildings, The University of Edinburgh, Edinburgh EH9 3JR, Scotland. Phone: 44 131 650 7092. Fax: 44 131 650 7040. E-mail:
d.tollervey{at}ed.ac.uk.
Present address: Department of Chemistry and Biochemistry,
University of California, Los Angeles, CA 90095-1569.
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