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Trends in Biochemical Sciences Jul 2023The mitochondrial ribosome (mitoribosome) is a multicomponent machine that has unique structural features. Biogenesis of the human mitoribosome includes correct... (Review)
Review
The mitochondrial ribosome (mitoribosome) is a multicomponent machine that has unique structural features. Biogenesis of the human mitoribosome includes correct maturation and folding of the mitochondria-encoded RNA components (12S and 16S mt-rRNAs, and mt-tRNAVal) and their assembly together with 82 nucleus-encoded mitoribosomal proteins. This complex process requires the coordinated action of multiple assembly factors. Recent advances in single-particle cryo-electron microscopy (cryo-EM) have provided detailed insights into the specific functions of several mitoribosome assembly factors and have defined their timing. In this review we summarize mitoribosomal small (mtSSU) and large subunit (mtLSU) biogenesis based on structural findings, and we discuss potential crosstalk between mtSSU and mtLSU assembly pathways as well as coordination between mitoribosome biogenesis and other processes involved in mitochondrial gene expression.
Topics: Humans; Cryoelectron Microscopy; Mitochondrial Ribosomes; RNA, Ribosomal, 16S; Mitochondrial Proteins; Ribosomal Proteins
PubMed: 37169615
DOI: 10.1016/j.tibs.2023.04.002 -
Nucleic Acids Research Sep 2023Archaeal ribosomes have many domain-specific features; however, our understanding of these structures is limited. We present 10 cryo-electron microscopy (cryo-EM)...
Archaeal ribosomes have many domain-specific features; however, our understanding of these structures is limited. We present 10 cryo-electron microscopy (cryo-EM) structures of the archaeal ribosome from crenarchaeota Sulfolobus acidocaldarius (Sac) at 2.7-5.7 Å resolution. We observed unstable conformations of H68 and h44 of ribosomal RNA (rRNA) in the subunit structures, which may interfere with subunit association. These subunit structures provided models for 12 rRNA expansion segments and 3 novel r-proteins. Furthermore, the 50S-aRF1 complex structure showed the unique domain orientation of aRF1, possibly explaining P-site transfer RNA (tRNA) release after translation termination. Sac 70S complexes were captured in seven distinct steps of the tRNA translocation reaction, confirming conserved structural features during archaeal ribosome translocation. In aEF2-engaged 70S ribosome complexes, 3D classification of cryo-EM data based on 30S head domain identified two new translocation intermediates with 30S head domain tilted 5-6° enabling its disengagement from the translocated tRNA and its release post-translocation. Additionally, we observed conformational changes to aEF2 during ribosome binding and switching from three different states. Our structural and biochemical data provide new insights into archaeal translation and ribosome translocation.
Topics: Cryoelectron Microscopy; Ribosomal Proteins; Ribosomes; RNA, Ribosomal; RNA, Transfer; Sulfolobus acidocaldarius
PubMed: 37604686
DOI: 10.1093/nar/gkad661 -
Current Opinion in Structural Biology Feb 2024During protein synthesis, the growing nascent peptide chain moves inside the polypeptide exit tunnel of the ribosome from the peptidyl transferase center towards the... (Review)
Review
During protein synthesis, the growing nascent peptide chain moves inside the polypeptide exit tunnel of the ribosome from the peptidyl transferase center towards the exit port where it emerges into the cytoplasm. The ribosome defines the unique energy landscape of the pioneering round of protein folding. The spatial confinement and the interactions of the nascent peptide with the tunnel walls facilitate formation of secondary structures, such as α-helices. The vectorial nature of protein folding inside the tunnel favors local intra- and inter-molecular interactions, thereby inducing cotranslational folding intermediates that do not form upon protein refolding in solution. Tertiary structures start to fold in the lower part of the tunnel, where interactions with the ribosome destabilize native protein folds. The present review summarizes the recent progress in understanding the driving forces of nascent protein folding inside the tunnel and at the surface of the ribosome.
Topics: Protein Folding; Ribosomes; Protein Biosynthesis; Proteins; Peptides
PubMed: 38071940
DOI: 10.1016/j.sbi.2023.102740 -
Clinical Epigenetics Jan 2024Periodontitis is a highly prevalent oral disease characterized by bacterium-induced periodontal inflammation and alveolar bone destruction. Osteoblast function is...
BACKGROUND
Periodontitis is a highly prevalent oral disease characterized by bacterium-induced periodontal inflammation and alveolar bone destruction. Osteoblast function is impaired in periodontitis with a global proteome change. METTL3 is the pivotal methyltransferase of N-methyladenosine (mA) that is recently proved to exert a crucial role in osteoblast differentiation. This study aims to investigate the role of METTL3 in osteoblast ribosome biogenesis in periodontitis progression.
RESULTS
METTL3 was knocked down in osteoblasts, and the downregulated genes were enriched in ribosome and translation. METTL3 knockdown inhibited ribosome biogenesis and oxidative phosphorylation in LPS-stimulated osteoblasts, whereas METTL3 overexpression facilitated ribosomal and mitochondrial function. Mechanistically, METTL3 mediated osteoblast biological behaviors by activating Wnt/β-catenin/c-Myc signaling. METTL3 depletion enhanced the mRNA expression and stability of Dkk3 and Sostdc1 via YTHDF2. In periodontitis mice, METTL3 inhibitor SAH promoted alveolar bone loss and local inflammatory status, which were partially rescued by Wnt/β-catenin pathway activator CHIR-99021 HCl.
CONCLUSIONS
METTL3 promoted ribosome biogenesis and oxidative phosphorylation by activating Wnt/β-catenin/c-Myc signaling in LPS-treated osteoblasts and alleviated the inflammatory alveolar bone destruction in periodontitis mice.
Topics: Animals; Mice; beta Catenin; DNA Methylation; Lipopolysaccharides; Methyltransferases; Osteoblasts; Periodontitis; Proto-Oncogene Proteins c-myc; Signal Transduction
PubMed: 38267969
DOI: 10.1186/s13148-024-01628-8 -
Science Advances Jun 2023Stem cells in many systems, including germline stem cells (GSCs), increase ribosome biogenesis and translation during terminal differentiation. Here, we show that the...
Stem cells in many systems, including germline stem cells (GSCs), increase ribosome biogenesis and translation during terminal differentiation. Here, we show that the H/ACA small nuclear ribonucleoprotein (snRNP) complex that promotes pseudouridylation of ribosomal RNA (rRNA) and ribosome biogenesis is required for oocyte specification. Reducing ribosome levels during differentiation decreased the translation of a subset of messenger RNAs that are enriched for CAG trinucleotide repeats and encode polyglutamine-containing proteins, including differentiation factors such as RNA-binding Fox protein 1. Moreover, ribosomes were enriched at CAG repeats within transcripts during oogenesis. Increasing target of rapamycin (TOR) activity to elevate ribosome levels in H/ACA snRNP complex-depleted germlines suppressed the GSC differentiation defects, whereas germlines treated with the TOR inhibitor rapamycin had reduced levels of polyglutamine-containing proteins. Thus, ribosome biogenesis and ribosome levels can control stem cell differentiation via selective translation of CAG repeat-containing transcripts.
Topics: Ribonucleoproteins, Small Nuclear; Ribosomes; RNA, Ribosomal; Proteins; Sirolimus
PubMed: 37343092
DOI: 10.1126/sciadv.ade5492 -
Proceedings of the National Academy of... Oct 2023Many mammalian proteins have circadian cycles of production and degradation, and many of these rhythms are altered posttranscriptionally. We used ribosome profiling to...
Many mammalian proteins have circadian cycles of production and degradation, and many of these rhythms are altered posttranscriptionally. We used ribosome profiling to examine posttranscriptional control of circadian rhythms by quantifying RNA translation in the liver over a 24-h period from circadian-entrained mice transferred to constant darkness conditions and by comparing ribosome binding levels to protein levels for 16 circadian proteins. We observed large differences in ribosome binding levels compared to protein levels, and we observed delays between peak ribosome binding and peak protein abundance. We found extensive binding of ribosomes to upstream open reading frames (uORFs) in circadian mRNAs, including the core clock gene . An increase in the number of uORFs in the 5'UTR was associated with a decrease in ribosome binding in the main coding sequence and a reduction in expression of synthetic reporter constructs. Mutation of the uORF increased luciferase and fluorescence reporter expression in 3T3 cells and increased luciferase expression in PER2:LUC MEF cells. Mutation of the uORF in mice increased mRNA expression, enhanced ribosome binding on , and reduced total sleep time compared to that in wild-type mice. These results suggest that uORFs affect mRNA posttranscriptionally, which can impact physiological rhythms and sleep.
Topics: Animals; Mice; Circadian Rhythm; Luciferases; Open Reading Frames; Ribosome Profiling; RNA, Messenger; Sleep; Period Circadian Proteins
PubMed: 37769257
DOI: 10.1073/pnas.2214636120 -
Trends in Biochemical Sciences Feb 2024Ribosomes interact with a variety of different protein biogenesis factors that guide newly synthesized proteins to their native 3D shapes and cellular localization.... (Review)
Review
Ribosomes interact with a variety of different protein biogenesis factors that guide newly synthesized proteins to their native 3D shapes and cellular localization. Depending on the type of translated substrate, a distinct set of cotranslational factors must interact with the ribosome in a timely and coordinated manner to ensure proper protein biogenesis. While cytonuclear proteins require cotranslational maturation and folding factors, secretory proteins must be maintained in an unfolded state and processed cotranslationally by transport and membrane translocation factors. Here we explore the specific cotranslational processing steps for cytonuclear, secretory, and membrane proteins in eukaryotes and then discuss how the nascent polypeptide-associated complex (NAC) cotranslationally sorts these proteins into the correct protein biogenesis pathway.
Topics: Ribosomes; Protein Biosynthesis; Protein Transport; Membrane Proteins; Saccharomyces cerevisiae
PubMed: 37919225
DOI: 10.1016/j.tibs.2023.10.003 -
Nature Structural & Molecular Biology Aug 2023The 5S ribonucleoprotein (RNP) is assembled from its three components (5S rRNA, Rpl5/uL18 and Rpl11/uL5) before being incorporated into the pre-60S subunit. However,...
The 5S ribonucleoprotein (RNP) is assembled from its three components (5S rRNA, Rpl5/uL18 and Rpl11/uL5) before being incorporated into the pre-60S subunit. However, when ribosome synthesis is disturbed, a free 5S RNP can enter the MDM2-p53 pathway to regulate cell cycle and apoptotic signaling. Here we reconstitute and determine the cryo-electron microscopy structure of the conserved hexameric 5S RNP with fungal or human factors. This reveals how the nascent 5S rRNA associates with the initial nuclear import complex Syo1-uL18-uL5 and, upon further recruitment of the nucleolar factors Rpf2 and Rrs1, develops into the 5S RNP precursor that can assemble into the pre-ribosome. In addition, we elucidate the structure of another 5S RNP intermediate, carrying the human ubiquitin ligase Mdm2, which unravels how this enzyme can be sequestered from its target substrate p53. Our data provide molecular insight into how the 5S RNP can mediate between ribosome biogenesis and cell proliferation.
Topics: Humans; RNA, Ribosomal, 5S; Tumor Suppressor Protein p53; Cryoelectron Microscopy; Ribosomal Proteins; Ribonucleoproteins; Ribosomes; Proto-Oncogene Proteins c-mdm2
PubMed: 37291423
DOI: 10.1038/s41594-023-01006-7 -
Molecular & Cellular Proteomics : MCP Sep 2023Ribosome profiling (Ribo-Seq) has proven transformative for our understanding of the human genome and proteome by illuminating thousands of noncanonical sites of...
Ribosome profiling (Ribo-Seq) has proven transformative for our understanding of the human genome and proteome by illuminating thousands of noncanonical sites of ribosome translation outside the currently annotated coding sequences (CDSs). A conservative estimate suggests that at least 7000 noncanonical ORFs are translated, which, at first glance, has the potential to expand the number of human protein CDSs by 30%, from ∼19,500 annotated CDSs to over 26,000 annotated CDSs. Yet, additional scrutiny of these ORFs has raised numerous questions about what fraction of them truly produce a protein product and what fraction of those can be understood as proteins according to conventional understanding of the term. Adding further complication is the fact that published estimates of noncanonical ORFs vary widely by around 30-fold, from several thousand to several hundred thousand. The summation of this research has left the genomics and proteomics communities both excited by the prospect of new coding regions in the human genome but searching for guidance on how to proceed. Here, we discuss the current state of noncanonical ORF research, databases, and interpretation, focusing on how to assess whether a given ORF can be said to be "protein coding."
Topics: Humans; Proteome; Protein Biosynthesis; Proteomics; Ribosome Profiling; Ribosomes; Open Reading Frames
PubMed: 37572790
DOI: 10.1016/j.mcpro.2023.100631