Small nucleolar RNAs (snoRNAs) are one of the most ancient and numerous families of non-protein-coding RNAs (ncRNAs). The main function of snoRNAs - to guide site-specific rRNA modification - is the same in Archaea and all eukaryotic lineages. In contrast, as revealed by recent genomic and RNomic studies, their genomic organization and expression strategies are the most varied. Seemingly snoRNA coding units have adopted, in the course of evolution, all the possible ways of being transcribed, thus providing a unique paradigm of gene expression flexibility. By focusing on representative fungal, plant and animal genomes, we review here all the documented types of snoRNA gene organization and expression, and we provide a comprehensive account of snoRNA expressional freedom by precisely estimating the frequency, in each genome, of each type of genomic organization. We finally discuss the relevance of snoRNA genomic studies for our general understanding of ncRNA family evolution and expression in eukaryotes.
Defects in ribosome biogenesis trigger stress response pathways, which perturb cell proliferation and differentiation in several genetic diseases. In Diamond–Blackfan anemia (DBA), a congenital erythroblastopenia, mutations in ribosomal protein genes often interfere with the processing of the internal transcribed spacer 1 (ITS1), the mechanism of which remains elusive in human cells. Using loss-of-function experiments and extensive RNA analysis, we have defined the precise position of the endonucleolytic cleavage E in the ITS1, which generates the 18S-E intermediate, the last precursor to the 18S rRNA. Unexpectedly, this cleavage is followed by 3′–5′ exonucleolytic trimming of the 18S-E precursor during nuclear export of the pre-40S particle, which sets a new mechanism for 18S rRNA formation clearly different from that established in yeast. In addition, cleavage at site E is also followed by 5′–3′ exonucleolytic trimming of the ITS1 by exonuclease XRN2. Perturbation of this step on knockdown of the large subunit ribosomal protein RPL26, which was recently associated to DBA, reveals the putative role of a highly conserved cis-acting sequence in ITS1 processing. These data cast new light on the original mechanism of ITS1 elimination in human cells and provide a mechanistic framework to further study the interplay of DBA-linked ribosomal proteins in this process.
Small nucleolar RNAs (snoRNAs) play a key role in ribosomal RNA biogenesis, yet factors controlling their expression are unknown. We found that the majority of Saccharomyces snoRNA promoters display an aRCCCTaa sequence motif at the upstream border of a TATA-containing nucleosome-free region. Genome-wide ChIP-seq analysis showed that these motifs are bound by Tbf1, a telomere-binding protein known to recognize mammalian-like T(2)AG(3) repeats at subtelomeric regions. Tbf1 has over 100 additional promoter targets, including several other genes involved in ribosome biogenesis and the TBF1 gene itself. Tbf1 is required for full snoRNA expression, yet it does not influence nucleosome positioning at snoRNA promoters. In contrast, Tbf1 contributes to nucleosome exclusion at non-snoRNA promoters, where it selectively colocalizes with the Tbf1-interacting zinc-finger proteins Vid22 and Ygr071c. Our data show that, besides the ribosomal protein gene regulator Rap1, a second telomere-binding protein also functions as a transcriptional regulator linked to yeast ribosome biogenesis.
Long-term cultures under hypoxic conditions have been demonstrated to maintain the phenotype of mesenchymal stromal/stem cells (MSCs) and to prevent the emergence of senescence. According to several studies, hypoxia has frequently been reported to drive genomic instability in cancer cells and in MSCs by hindering the DNA damage response and DNA repair. Thus, we evaluated the occurrence of DNA damage and repair events during the ex vivo expansion of clinical-grade adipose-derived stromal cells (ADSCs) and bone marrow (BM)-derived MSCs cultured with platelet lysate under 21% (normoxia) or 1% (hypoxia) O 2 conditions. Hypoxia did not impair cell survival after DNA damage, regardless of MSC origin. However, ADSCs, unlike BMMSCs, displayed altered cH2AX signaling and increased ubiquitylated cH2AX levels under hypoxic conditions, indicating an impaired resolution of DNA damage-induced foci. Moreover, hypoxia specifically promoted BM-MSC DNA integrity, with increased Ku80, TP53BP1, BRCA1, and RAD51 expression levels and more efficient nonhomologous end joining and homologous recombination repair. We further observed that hypoxia favored mtDNA stability and maintenance of differentiation potential after genotoxic stress. We conclude that long-term cultures under 1% O 2 were more suitable for BM-MSCs as suggested by improved genomic stability compared with ADSCs. STEM CELLS 2015;33:3608-3620 SIGNIFICANCE STATEMENTHuman mesenchymal stem cells (MSCs) feature differentiation capacities, immunomodulatory properties and therefore represent a great potential for medicine. In vitro culture allows their amplification for suitable clinical uses. Hypoxia seems relevant for the prevention of senescence and loss of MSC features. However, hypoxia is shown to promote cancer or DNA damage response (DDR) and repair failures. We demonstrate that long-term culture of MSCs with platelet lysate in hypoxia does not necessarily inhibit DDR and DNA repair. Here, bone marrowMSCs conserved or rather improved their DDR and DNA repair (NHEJ, HR), mtDNA stability in hypoxia compared to adipose-derived stem cells. Moreover, the ubiquitylation status of gH2AX between BM-MSCs and ADSCs might be responsible for changes in their DNA repair activities. Appropriate oxygen tension seems crucial to preserve the MSCs genomic stability according to their origins.
The Saccharomyces cerevisiae SNR52 gene is unique among the snoRNA coding genes in being transcribed by RNA polymerase III. The primary transcript of SNR52 is a 250-nucleotide precursor RNA from which a long leader sequence is cleaved to generate the mature snR52 RNA. We found that the box A and box B sequence elements in the leader region are both required for the in vivo accumulation of the snoRNA. As expected box B, but not box A, was absolutely required for stable TFIIIC, yet in vitro. Surprisingly, however, the box B was found to be largely dispensable for in vitro transcription of SNR52, whereas the box A-mutated template effectively recruited TFIIIB; yet it was transcriptionally inactive. Even in the complete absence of box B and both upstream TATA-like and T-rich elements, the box A still directed efficient, TFIIIC-dependent transcription. Box B-independent transcription was also observed for two members of the tRNA Asn (GTT) gene family, but not for two tRNA Pro (AGG) gene copies. Fully recombinant TFIIIC supported box B-independent transcription of both SNR52 and tRNA Asn genes, but only in the presence of TFIIIB reconstituted with a crude B؆ fraction. Non-TFIIIB component(s) in this fraction were also required for transcription of wild-type SNR52. Transcription of the box B-less tRNA Asn genes was strongly influenced by their 5-flanking regions, and it was stimulated by TBP and Brf1 proteins synergistically. The box A can thus be viewed as a core TFIIICinteracting element that, assisted by upstream TFIIIB-DNA contacts, is sufficient to promote class III gene transcription.RNA polymerase (Pol) 3 III synthesizes tRNA, 5 S rRNA, and a variety of other types of small nuclear and cytoplasmic RNAs. In general, the transcription of class III genes is under the control of internal control regions (ICR) characterized by discontinuous structures, with essential boxes separated by non-essential nucleotides (1). In the case of tRNA and 5 S rRNA genes, the ICRs are highly conserved and comprise the binding sites for the general transcription factor TFIIIC (box A and box B) and for the 5 S-specific factor TFIIIA (box C). Once assembled, TFIIIC recruits TFIIIB upstream of the transcription start site (TSS); TFIIIB in turn recruits Pol III for transcription initiation. The strong conservation of the ICRs likely reflects their dual function as both nucleation sites for transcription complex assembly and key determinants of tRNA and 5 S rRNA structure. An indication of the constraints imposed on ICR by their overlapping structural and functional roles comes from the variability of promoter organizations displayed by the minority of class III genes not coding for tRNA and 5 S rRNA. In some of these genes, the TFIIIC-interacting control regions (box A and box B) have been maintained within the transcribed region, and adapted to the structure of the small RNA without losing the transcriptional function. For example, in the Saccharomyces cerevisiae SCR1 gene, coding for the 7SL RNA, box A and box B are both intragenic an...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.