Phosphorylation of the Saccharomyces cerevisiae La protein does not appear to be required for its functions in tRNA maturation and nascent RNA stabilization
(2001) In RNA 7(11). p.1589-1602- Abstract
An abundant nuclear phosphoprotein, the La autoantigen, is the first protein to bind all newly synthesized RNA polymerase III transcripts. Binding by the La protein to the 3′ ends of these RNAs stabilizes the nascent transcripts from exonucleolytic degradation. In the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, the La protein is required for the normal pathway of tRNA maturation. Experiments in which the human protein was expressed in S. pombe have suggested that phosphorylation of the La protein regulates tRNA maturation. To dissect the role of phosphorylation in La protein function, we used mass spectrometry to identify three sites of serine phosphorylation in the S. cerevisiae La protein... (More)
An abundant nuclear phosphoprotein, the La autoantigen, is the first protein to bind all newly synthesized RNA polymerase III transcripts. Binding by the La protein to the 3′ ends of these RNAs stabilizes the nascent transcripts from exonucleolytic degradation. In the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, the La protein is required for the normal pathway of tRNA maturation. Experiments in which the human protein was expressed in S. pombe have suggested that phosphorylation of the La protein regulates tRNA maturation. To dissect the role of phosphorylation in La protein function, we used mass spectrometry to identify three sites of serine phosphorylation in the S. cerevisiae La protein Lhp1p. Mutant versions of Lhp1p, in which each of the serines was mutated to alanine, were expressed in yeast cells lacking Lhp1p. Using two-dimensional gel electrophoresis, we determined that we had identified and mutated all major sites of phosphorylation in Lhp1p. Lhp1p lacking all three phosphorylation sites was functional in several yeast strains that require Lhp1p for growth. Northern blotting revealed no effects of Lhp1p phosphorylation status on either pre-tRNA maturation or stabilization of nascent RNAs. Both wild-type and mutant Lhp1 proteins localized to both nucleoplasm and nucleoli, demonstrating that phosphorylation does not affect subcellular location. Thus, although La proteins from yeast to humans are phosphoproteins, phosphorylation does not appear to be required for any of the identified functions of the S. cerevisiae protein.
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- author
- Long, Katherine S. ; Cedervall, Tommy LU ; Walch-Solimena, Christiane ; Noe, Dennis A. ; Huddleston, Michael J. ; Annan, Roland S. and Wolin, Sandra L.
- publishing date
- 2001
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Lhp1p, Mass spectrometry, Nucleolus, RNA-binding proteins, RNA-protein interactions
- in
- RNA
- volume
- 7
- issue
- 11
- pages
- 14 pages
- publisher
- Cold Spring Harbor Laboratory Press (CSHL)
- external identifiers
-
- scopus:0035163788
- pmid:11720288
- ISSN
- 1355-8382
- language
- English
- LU publication?
- no
- id
- de1a4987-4354-4f48-8214-dba744459f55
- alternative location
- https://www.cambridge.org/core/journals/rna/article/abs/phosphorylation-of-the-saccharomyces-cerevisiae-la-protein-does-not-appear-to-be-required-for-its-functions-in-trna-maturation-and-nascent-rna-stabilization/28F169B9F1406BA481683734750861C9
- date added to LUP
- 2022-02-10 08:16:18
- date last changed
- 2024-01-07 08:06:14
@article{de1a4987-4354-4f48-8214-dba744459f55,
abstract = {{<p>An abundant nuclear phosphoprotein, the La autoantigen, is the first protein to bind all newly synthesized RNA polymerase III transcripts. Binding by the La protein to the 3′ ends of these RNAs stabilizes the nascent transcripts from exonucleolytic degradation. In the yeasts <i>Saccharomyces cerevisiae</i> and <i>Schizosaccharomyces pombe</i>, the La protein is required for the normal pathway of tRNA maturation. Experiments in which the human protein was expressed in <i>S. pombe</i> have suggested that phosphorylation of the La protein regulates tRNA maturation. To dissect the role of phosphorylation in La protein function, we used mass spectrometry to identify three sites of serine phosphorylation in the <i>S. cerevisiae</i> La protein Lhp1p. Mutant versions of Lhp1p, in which each of the serines was mutated to alanine, were expressed in yeast cells lacking Lhp1p. Using two-dimensional gel electrophoresis, we determined that we had identified and mutated all major sites of phosphorylation in Lhp1p. Lhp1p lacking all three phosphorylation sites was functional in several yeast strains that require Lhp1p for growth. Northern blotting revealed no effects of Lhp1p phosphorylation status on either pre-tRNA maturation or stabilization of nascent RNAs. Both wild-type and mutant Lhp1 proteins localized to both nucleoplasm and nucleoli, demonstrating that phosphorylation does not affect subcellular location. Thus, although La proteins from yeast to humans are phosphoproteins, phosphorylation does not appear to be required for any of the identified functions of the <i>S. cerevisiae</i> protein.</p>}},
author = {{Long, Katherine S. and Cedervall, Tommy and Walch-Solimena, Christiane and Noe, Dennis A. and Huddleston, Michael J. and Annan, Roland S. and Wolin, Sandra L.}},
issn = {{1355-8382}},
keywords = {{Lhp1p; Mass spectrometry; Nucleolus; RNA-binding proteins; RNA-protein interactions}},
language = {{eng}},
number = {{11}},
pages = {{1589--1602}},
publisher = {{Cold Spring Harbor Laboratory Press (CSHL)}},
series = {{RNA}},
title = {{Phosphorylation of the <i>Saccharomyces cerevisiae</i> La protein does not appear to be required for its functions in tRNA maturation and nascent RNA stabilization}},
url = {{https://www.cambridge.org/core/journals/rna/article/abs/phosphorylation-of-the-saccharomyces-cerevisiae-la-protein-does-not-appear-to-be-required-for-its-functions-in-trna-maturation-and-nascent-rna-stabilization/28F169B9F1406BA481683734750861C9}},
volume = {{7}},
year = {{2001}},
}