The La protein functions redundantly with tRNA modification enzymes to ensure tRNA structural stability

Copela, Laura A.; Chakshusmathi, Ghadiyaram; Sherrer, R. Lynn; Wolin, Sandra L.

doi:10.1261/rna.2307206

Cited by 63 publications

(79 citation statements)

References 53 publications

(71 reference statements)

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“…For example, Lhp1p becomes required for accumulation of the U6 spliceosomal snRNA when yeast contain mutations in the Lsm2-Lsm8 proteins that are components of the mature RNP (4,5). For pre-tRNAs, both modifications and binding to subsequent proteins such as synthetases likely contribute to maintaining correct folds in vivo, because Lhp1p is important for growth when yeast contain mutations in either the arginyl-tRNA synthetase or the tRNA modifying enzyme TRM1 (29). Consistent with a redundant function in stabilizing tRNA structure, Trm1p catalyzes dimethylguanosine formation at position 26 in many tRNAs, a modification that prevents formation of alternate anticodon stems (30).…”

Section: Discussionmentioning

confidence: 99%

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An intrinsically disordered C terminus allows the La protein to assist the biogenesis of diverse noncoding RNA precursors

Kucera

Hodsdon

Wolin

2011

Proc. Natl. Acad. Sci. U.S.A.

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The La protein binds the 3′ ends of many newly synthesized noncoding RNAs, protecting these RNAs from nucleases and influencing folding, maturation, and ribonucleoprotein assembly. Although 3′ end binding by La involves the N-terminal La domain and adjacent RNA recognition motif (RRM), the mechanisms by which La stabilizes diverse RNAs from nucleases and assists subsequent events in their biogenesis are unknown. Here we report that a conserved feature of La proteins, an intrinsically disordered C terminus, is required for the accumulation of certain noncoding RNA precursors and for the role of the Saccharomyces cerevisiae La protein Lhp1p in assisting formation of correctly folded pre-tRNA anticodon stems in vivo. Footprinting experiments using purified Lhp1p reveal that the C terminus is required to protect a pre-tRNA anticodon stem from chemical modification. Although the C terminus of Lhp1p is hypersensitive to proteases in vitro, it becomes protease-resistant upon binding pre-tRNAs, U6 RNA, or pre-5S rRNA. Thus, while high affinity binding to 3′ ends requires the La domain and RRM, a conformationally flexible C terminus allows La to interact productively with a diversity of noncoding RNA precursors. We propose that intrinsically disordered domains adjacent to well characterized RNA-binding motifs in other promiscuous RNA-binding proteins may similarly contribute to the ability of these proteins to influence the cellular fates of multiple distinct RNA targets.RNA-binding protein | unstructured domain | noncoding RNA biogenesis M any RNA-binding proteins interact with multiple ligands, influencing the fate of the RNAs and in some cases altering structure. For example, the pre-mRNA binding protein hnRNP A1 and the mRNA decay mediator KSRP also bind miRNA precursors to promote their maturation (1, 2). Similarly, the Y-box protein YB-1, a component of cytoplasmic mRNPs, functions in both pre-mRNA splicing and translational repression of some mRNAs (3). Although most of these proteins contain well characterized RNA-binding motifs, the mechanisms by which they interact with diverse targets and influence their conformations and outcomes are not understood.One such protein, called La, is a nuclear phosphoprotein whose best characterized role is to assist noncoding RNA biogenesis. La binds the UUU OH that is the initial end of all RNA polymerase III transcripts and protects the RNAs from exonucleases (4, 5). In budding yeast, La also binds processing intermediates of RNA polymerase II-transcribed small nuclear and nucleolar RNAs that end in UUU OH (6-8). Because most La-bound RNAs undergo 3′ maturation, La is usually not bound to the mature RNAs. Binding by La results in diverse consequences. For pretRNAs, La binding favors endonucleolytic removal of the trailer (9) and is required for wild-type levels of some tRNAs (10). For the yeast U4 snRNA, the La-bound precursor preferentially associates with the core Sm proteins in vitro, suggesting that La assists snRNP formation by protecting the favored substrate for Sm prot...

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Section: Discussionmentioning

confidence: 99%

“…Monoclonal anti-Nop1p was a gift of J. Aris (University of Florida, Gainesville). Northern blotting was as described (29). To examine aminoacylation, RNA was extracted at low pH and fractionated in acidic polyacrylamide gels (33).…”

Section: Methodsmentioning

confidence: 99%

An intrinsically disordered C terminus allows the La protein to assist the biogenesis of diverse noncoding RNA precursors

Kucera

Hodsdon

Wolin

2011

Proc. Natl. Acad. Sci. U.S.A.

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“…Analyses of the mutant strains revealed that La stabilizes nascent U6 snRNAs when cells contain mutations in components of the Lsm2-Lsm8 ring (Pannone et al 1998(Pannone et al , 2001) and that La assists assembly of the U4 snRNA into the U4/U6 snRNP when cells contain a mutation in the snRNP core protein Smd1p (Xue et al 2000). La is also important for growth when cells contain mutations in the arginyl tRNA synthetase or either of two tRNA modification enzymes, Trm1p and Trm61p (Calvo et al 1999;Copela et al 2006). Taken together, these experiments suggest that La functions redundantly with other proteins that contact nascent RNAs to stabilize these RNAs and assist their assembly into functional RNPs.…”

Section: La Stabilizes Nascent Noncoding Rnas and Assists Pre-trna Fomentioning

confidence: 99%

“…In addition to the requirement for La, the trr4-1 mutation causes a fraction of the mutant tRNA to be unstable. Degradation of the mutant tRNA does not require the Trf4 polymerase (Copela et al 2006). (B) Methylation of adenosine 58 is required for stable accumulation of tRNA i Met .…”

Section: Sm-like Protein Rings May Facilitate Rna-rna Interactionsmentioning

confidence: 99%

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Molecular Chaperones and Quality Control in Noncoding RNA Biogenesis

Wolin¹,

Wurtmann²

2006

Cold Spring Harbor Symposia on Quantitative Biology

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Although noncoding RNAs have critical roles in all cells, both the mechanisms by which these RNAs fold into functional structures and the quality control pathways that monitor correct folding are only beginning to be elucidated. Here, we discuss several proteins that likely function as molecular chaperones for noncoding RNAs and review the existing knowledge on noncoding RNA quality control. One protein, the La protein, binds many nascent noncoding RNAs in eukaryotes and is required for efficient folding of certain pre-tRNAs. In prokaryotes, the Sm-like protein Hfq is required for the function of many noncoding RNAs. Recent work in bacteria and yeast has revealed the existence of quality control systems involving polyadenylation of unstable noncoding RNAs followed by exonucleolytic degradation. In addition, the Ro protein, which is present in many animal cells and also certain bacteria, binds misfolded noncoding RNAs and is proposed to function in RNA quality control.

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The RNA‐binding protein La/SSB associates with radiation‐induced DNA double‐strand breaks in lung cancer cell lines

Staudacher

Liapis

et al. 2021

Cancer Reports

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Background Platinum‐based chemotherapy and radiotherapy are standard treatments for non‐small cell lung cancer, which is the commonest, most lethal cancer worldwide. As a marker of treatment‐induced cancer cell death, we have developed a radiodiagnostic imaging antibody, which binds to La/SSB. La/SSB is an essential, ubiquitous ribonuclear protein, which is over expressed in cancer and plays a role in resistance to cancer therapies. Aim In this study, we examined radiation‐induced DNA double strand breaks (DSB) in lung cancer cell lines and examined whether La/SSB associated with these DSB. Method Three lung cancer lines (A549, H460 and LL2) were irradiated with different X‐ray doses or X‐radiated with a 5 Gy dose and examined at different time‐points post‐irradiation for DNA DSB in the form of γ‐H2AX and Rad51 foci. Using fluorescence microscopy, we examined whether La/SSB and γ‐H2AX co‐localise and performed proximity ligation assay (PLA) and co‐immunoprecipitation to confirm the interaction of these proteins. Results We found that the radio‐resistant A549 cell line compared to the radio‐sensitive H460 cell line showed faster resolution of radiation‐induced γ‐H2AX foci over time. Conversely, we found more co‐localised γ‐H2AX and La/SSB foci by PLA in irradiated A549 cells. Conclusion The co‐localisation of La/SSB with radiation‐induced DNA breaks suggests a role of La/SSB in DNA repair, however further experimentation is required to validate this.

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The La protein functions redundantly with tRNA modification enzymes to ensure tRNA structural stability

Cited by 63 publications

References 53 publications

An intrinsically disordered C terminus allows the La protein to assist the biogenesis of diverse noncoding RNA precursors

An intrinsically disordered C terminus allows the La protein to assist the biogenesis of diverse noncoding RNA precursors

Molecular Chaperones and Quality Control in Noncoding RNA Biogenesis

The RNA‐binding protein La/SSB associates with radiation‐induced DNA double‐strand breaks in lung cancer cell lines

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