Precursor molecules of both human 5S ribosomal RNA and transfer RNAs are bound by a cellular protein reactive with anti-La Lupus antibodies

doi:10.1016/0092-8674(82)90099-X

Cell

Volume 29, Issue 1, May 1982, Pages 149-159

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Abstract

The small ribonucleoproteins recognized by anti-La autoantibodies contain a heterogeneous mixture of small RNAs from uninfected mammalian cells. The identity of many of these has now been established by the discovery of precursor forms of 5S rRNA and of certain tRNAs among La RNAs from HeLa cells. The small fraction of 5S rRNA molecules that exist as La ribonucleoproteins in vivo possess 1 or 2 additional U residues at their 3′ ends. Such 5S molecules bound to the La protein have also been identified with in vitro nuclear transcription systems. Pulse-chase experiments performed both in vivo and in vitro support the idea that most newly synthesized 5S rRNA molecules are transiently associated with the La protein. Cell extracts contain a processing activity that converts longer in vitro-synthesized 5S RNA transcripts into molecules of mature size. The presence of in vivo tRNA precursors in the heterogeneous mixture of La RNAs is demonstrated by the identification of precursor forms of five different specific tRNAs (Met_i, Asp, Gly, Glu, Asn). After in vitro transcription of a tRNA gene (tRNA_i^Met), only products the size of precursor molecules are precipitable by anti-La antibodies. The realization that virtually every known RNA polymerase III product associates at least initially with the La antigen suggests that this protein plays an essential role in the synthesis or maturation of all class III transcripts.

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Cited by (256)

Crosstalk between the tRNA methyltransferase Trm1 and RNA chaperone La influences eukaryotic tRNA maturation
2023, Journal of Biological Chemistry
tRNAs undergo an extensive maturation process involving posttranscriptional modifications often associated with tRNA structural stability and promoting the native fold. Impaired posttranscriptional modification has been linked to human disease, likely through defects in translation, mitochondrial function, and increased susceptibility to degradation by various tRNA decay pathways. More recently, evidence has emerged that bacterial tRNA modification enzymes can act as tRNA chaperones to guide tRNA folding in a manner independent from catalytic activity. Here, we provide evidence that the fission yeast tRNA methyltransferase Trm1, which dimethylates nuclear- and mitochondrial-encoded tRNAs at G26, can also promote tRNA functionality in the absence of catalysis. We show that WT and catalytic-dead Trm1 are active in an in vivo tRNA-mediated suppression assay and possess RNA strand annealing and dissociation activity in vitro, similar to previously characterized RNA chaperones. Trm1 and the RNA chaperone La have previously been proposed to function synergistically in promoting tRNA maturation, yet we surprisingly demonstrate that La binding to nascent pre-tRNAs decreases Trm1 tRNA dimethylation in vivo and in vitro. Collectively, these results support the hypothesis for tRNA modification enzymes that combine catalytic and noncatalytic activities to promote tRNA maturation, as well as expand our understanding of how La function can influence tRNA modification.
Ribosomal RNA Processing
2022, Encyclopedia of Cell Biology: Volume 1-6, Second Edition
Eukaryotic ribosomes are extraordinarily complex cellular machines that catalyze the peptide bond synthesis necessary for producing new proteins. Ribosomes are composed of protein and RNA components, but the catalytic activity is found within the RNA itself. Each ribosome contains four ribosomal RNAs (rRNAs) initially synthesized from two precursor genes. These precursors undergo a laborious maturation process to release the mature rRNAs. In this article, we will discuss the mechanistic details governing pre-rRNA processing, focusing on those processes in human and yeast cells.
The Autoantigen Repertoire and the Microbial RNP World
2021, Trends in Molecular Medicine
Citation Excerpt :
The targeted RNPs (Table 1) include SRP, RNase P, and spliceosomal snRNPs, such as the U1 snRNP that is recognized by both anti-nRNP (also termed anti-RNP and anti-U1 RNP) and anti-Sm autoantibodies [49–51]. Other targeted RNPs include complexes of the Ro60 protein and noncoding RNAs termed Y RNAs that are recognized by anti-Ro (also termed SSA) autoantibodies, and complexes between La protein and newly synthesized tRNAs or other noncoding RNAs that are recognized by anti-La (also termed SSB) autoantibodies [52–57]. Some autoantibodies are specific for particular diseases (Table 2).
Although autoimmunity and autoimmune disease (AID) are relatively common, the repertoire of autoantigens is paradoxically very limited. Highly enriched in this autoantigen repertoire are nucleic acids and their binding proteins, which together form large macromolecular structures. Most of these complexes are of ancient evolutionary origin, with homologs throughout multiple kingdoms of life. Why and if these nucleic acid–protein particles drive the development of autoimmunity remains unresolved. Recent advances in our understanding of the microbiome may provide clues about the origins of autoimmunity – and the particular puzzle of why the autoantigen repertoire is so particularly enriched in ribonucleoprotein particles (RNPs). We discuss the possibility that autoimmunity to some RNPs may arise from molecular mimicry to microbial orthologs.
Interactions between RNAP III transcription machinery and tRNA processing factors
2018, Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
Citation Excerpt :
While in wild-type yeast cells the 5′ leader cleavage precedes other processing steps, in cells that lack La protein, cells undergo a 3′ first pathway, indicating that in these cells, 5′ leader cleavage occurs only after the release of the transcripts by RNAP III [29]. Since the primary binding site of La is the 3′-U-OH of nascent tRNAs, it is apparent that stable La-tRNA interaction occur only after the release of transcript from RNAP III [75–77]. The apparent lack of co-transcriptional tRNA processing could be due to the inherent nature of RNAP III transcription.
Eukaryotes have at least three nuclear RNA polymerases to carry out transcription. While RNA polymerases I and II are responsible for ribosomal RNA transcription and messenger RNA transcription, respectively, RNA Polymerase III transcribes approximately up to 300 nt long noncoding RNAs, including tRNA. For all three RNAPs, the nascent transcripts generated undergo extensive post-transcriptional processing. Transcription of mRNAs by RNAP II and their processing are coupled with the aid of the C-terminal domain of the RNAP II. RNAP I transcription and the processing of its transcripts are co-localized to the nucleolus and to some extent, rRNA processing occurs co-transcriptionally. Here, I review the current evidence for the interaction between tRNA processing factors and RNA polymerase III. These interactions include the moonlighting functions of tRNA processing factors in RNAP III transcription and the indirect effect of tRNA transcription levels on tRNA modification machinery.
La involvement in tRNA and other RNA processing events including differences among yeast and other eukaryotes
2018, Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
Citation Excerpt :
The snRNAs bound to La, were found to be those synthesized by RNA polymerase (RNAP) III. RNAP III transcripts all share a common UUU-3′OH motif that represents the transcription termination signal for RNAP III, and the high-affinity, sequence-specific recognition site for La [3–6] (reviewed in [7]). The cloning and analysis of the cDNA encoding human La protein revealed this founding member of a class of RNA-binding proteins that contain sequence determinants defining an RNA recognition motif (RRM) [8–10].
The conserved nuclear RNA-binding factor known as La protein arose in an ancient eukaryote, phylogenetically associated with another eukaryotic hallmark, synthesis of tRNA by RNA polymerase III (RNAP III). Because 3′-oligo(U) is the sequence-specific signal for transcription termination by RNAP III as well as the high affinity binding site for La, the latter is linked to the intranuclear posttranscriptional processing of eukaryotic precursor-tRNAs. The pre-tRNA processing pathway must accommodate a variety of substrates that are destined for both common steps as well as tRNA-specific events. The order of intranuclear pre-tRNA processing steps is mediated in part by three activities derived from interaction with La protein: 3′-end protection from untimely decay by 3′ exonucleases, nuclear retention and chaperone activity that helps prevent pre-tRNA misfolding and mischanneling into offline pathways. A focus of this perspective will be on differences between yeast and mammals in the subcellular partitioning of pre-tRNA intermediates and differential interactions with La. We review how this is most relevant to pre-tRNA splicing which occurs in the cytoplasm of yeasts but in nuclei of higher eukaryotes. Also divergent is La architecture, comprised of three RNA-binding domains in organisms in all examined branches of the eukaryal tree except yeast, which have lost the C-terminal RNA recognition motif-2α (RRM2α) domain. We also review emerging data that suggest mammalian La interacts with nuclear pre-tRNA splicing intermediates and may impact this branch of the tRNA maturation pathway. Finally, because La is involved in intranuclear tRNA biogenesis we review relevant aspects of tRNA-associated neurodegenerative diseases. This article is part of a Special Issue entitled: SI: Regulation of tRNA synthesis and modification in physiological conditions and disease edited by Dr. Boguta Magdalena.
Adenovirus VA RNA: An essential pro-viral non-coding RNA
2016, Virus Research
Citation Excerpt :
VA RNAI is present in both the nucleus and cytoplasm of AdV-infected cells, but accumulates to high copy number in the cytoplasm late in infection. The La autoantigen is a protein recognized by the sera of patients with systematic lupus erythematosus, which binds to the 3′-end UUU-OH sequence of RNA Pol III transcripts, including tRNAs, pre-5S rRNA and VA RNAI (Lerner et al., 1981; Rinke and Steitz, 1982; Wolin and Cedervall, 2002). La binding facilitates proper maturation of cellular RNA Pol III transcripts, contributes to nuclear retention of small RNAs, facilitates formation of ribonucleoprotein complexes, and can promote translation of specific mRNAs (reviewed in Wolin and Cedervall, 2002).
Adenovirus (AdV) ‘virus-associated’ RNAs (VA RNAs) are exceptionally abundant (up to 10⁸ copies/cell), heterogeneous, non-coding RNA transcripts (∼150–200 nucleotides). The predominant species, VA RNA_I, is best recognized for its essential function in relieving the cellular anti-viral blockade of protein synthesis through inhibition of the double-stranded RNA-activated protein kinase (PKR). More recent evidence has revealed that VA RNAs also interfere with several other host cell processes, in part by virtue of the high level to which they accumulate. Following transcription by cellular RNA polymerase III, VA RNAs saturate the nuclear export protein Exportin 5 (Exp5) and the cellular endoribonculease Dicer, interfering with pre-micro (mi)RNA export and miRNA biogenesis, respectively. Dicer-processed VA RNA fragments are incorporated into the RNA-induced silencing complex (RISC) as ‘mivaRNAs’, where they may specifically target cellular genes. VA RNA_I also interacts with other innate immune proteins, including OAS1. While intact VA RNA_I has the paradoxical effect of activating OAS1, a non-natural VA RNA_I construct lacking the entire Terminal Stem has been reported to be a pseudoinhibitor of OAS1. Here, we show that a VA RNA_I construct corresponding to an authentic product of Dicer processing similarly fails to activate OAS1 but also retains only a modest level of inhibitory activity against PKR in contrast to the non-natural deletion construct. These findings underscore the complexity of the arms race between virus and host, and highlight the need for further exploration of the impact of VA RNA_I interactions with host defenses on the outcome of AdV infection beyond that of well-established PKR inhibition. Additional contributions of VA RNA_I heterogeneity resulting from variations in transcription initiation and termination to each of these functions remain open questions that are discussed here.

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^∗: Present address: Max Planck Institute for Molecular Genetics, Ihnestrasse 63–73, D-1000 Berlin 33, Federal Republic of Germany.

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ArticlePrecursor molecules of both human 5S ribosomal RNA and transfer RNAs are bound by a cellular protein reactive with anti-La Lupus antibodies

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Precursor molecules of both human 5S ribosomal RNA and transfer RNAs are bound by a cellular protein reactive with anti-La Lupus antibodies