bims-tricox Biomed News
on Translation, ribosomes and COX
Issue of 2022–10–02
nine papers selected by
Yash Verma, University of Delhi South Campus
  1. ANGEL2 phosphatase activity is required for non-canonical mitochondrial RNA processing.
  2. Human NOP2/NSUN1 regulates ribosome biogenesis through non-catalytic complex formation with box C/D snoRNPs.
  3. Visualizing translation dynamics at atomic detail inside a bacterial cell.
  4. A Tale of Water Molecules in the Ribosomal Peptidyl Transferase Reaction.
  5. On the role of ubiquinone in the proton translocation mechanism of respiratory complex I.
  6. Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3.
  7. Mass spectrometry analysis of affinity-purified cytoplasmic translation initiation complexes from human and fly cells.
  8. Promoting a new view of mitochondrial genome regulation.
  9. Recent advances in mitochondrial diseases: From molecular insights to therapeutic perspectives.
  1. Nat Commun. 2022 Sep 30. 13(1): 5750
    ANGEL2 phosphatase activity is required for non-canonical mitochondrial RNA processing.
    Paula Clemente, Javier Calvo-Garrido, Sarah F Pearce, Florian A Schober, Megumi Shigematsu, Stefan J Siira, Isabelle Laine, Henrik Spåhr, Christian Steinmetzger, Katja Petzold, Yohei Kirino, Rolf Wibom, Oliver Rackham, Aleksandra Filipovska, Joanna Rorbach, Christoph Freyer, Anna Wredenberg.
      Canonical RNA processing in mammalian mitochondria is defined by tRNAs acting as recognition sites for nucleases to release flanking transcripts. The relevant factors, their structures, and mechanism are well described, but not all mitochondrial transcripts are punctuated by tRNAs, and their mode of processing has remained unsolved. Using Drosophila and mouse models, we demonstrate that non-canonical processing results in the formation of 3' phosphates, and that phosphatase activity by the carbon catabolite repressor 4 domain-containing family member ANGEL2 is required for their hydrolysis. Furthermore, our data suggest that members of the FAST kinase domain-containing protein family are responsible for these 3' phosphates. Our results therefore propose a mechanism for non-canonical RNA processing in metazoan mitochondria, by identifying the role of ANGEL2.
    DOI:  https://doi.org/10.1038/s41467-022-33368-9
  2. Nucleic Acids Res. 2022 Sep 26. pii: gkac817. [Epub ahead of print]
    Human NOP2/NSUN1 regulates ribosome biogenesis through non-catalytic complex formation with box C/D snoRNPs.
    Han Liao, Anushri Gaur, Hunter McConie, Amirtha Shekar, Karen Wang, Jeffrey T Chang, Ghislain Breton, Catherine Denicourt.
      5-Methylcytosine (m5C) is a base modification broadly found on various RNAs in the human transcriptome. In eukaryotes, m5C is catalyzed by enzymes of the NSUN family composed of seven human members (NSUN1-7). NOP2/NSUN1 has been primarily characterized in budding yeast as an essential ribosome biogenesis factor required for the deposition of m5C on the 25S ribosomal RNA (rRNA). Although human NOP2/NSUN1 has been known to be an oncogene overexpressed in several types of cancer, its functions and substrates remain poorly characterized. Here, we used a miCLIP-seq approach to identify human NOP2/NSUN1 RNA substrates. Our analysis revealed that NOP2/NSUN1 catalyzes the deposition of m5C at position 4447 on the 28S rRNA. We also find that NOP2/NSUN1 binds to the 5'ETS region of the pre-rRNA transcript and regulates pre-rRNA processing through non-catalytic complex formation with box C/D snoRNAs. We provide evidence that NOP2/NSUN1 facilitates the recruitment of U3 and U8 snoRNAs to pre-90S ribosomal particles and their stable assembly into snoRNP complexes. Remarkably, expression of both WT and catalytically inactive NOP2/NSUN1 in knockdown background rescues the rRNA processing defects and the stable assembly of box C/D snoRNP complexes, suggesting that NOP2/NSUN1-mediated deposition of m5C on rRNA is not required for ribosome synthesis.
    DOI:  https://doi.org/10.1093/nar/gkac817
  3. Nature. 2022 Sep 28.
    Visualizing translation dynamics at atomic detail inside a bacterial cell.
    Liang Xue, Swantje Lenz, Maria Zimmermann-Kogadeeva, Dimitry Tegunov, Patrick Cramer, Peer Bork, Juri Rappsilber, Julia Mahamid.
      Translation is the fundamental process of protein synthesis and is catalysed by the ribosome in all living cells1. Here we use advances in cryo-electron tomography and sub-tomogram analysis2,3 to visualize the structural dynamics of translation inside the bacterium Mycoplasma pneumoniae. To interpret the functional states in detail, we first obtain a high-resolution in-cell average map of all translating ribosomes and build an atomic model for the M. pneumoniae ribosome that reveals distinct extensions of ribosomal proteins. Classification then resolves 13 ribosome states that differ in their conformation and composition. These recapitulate major states that were previously resolved in vitro, and reflect intermediates during active translation. On the basis of these states, we animate translation elongation inside native cells and show how antibiotics reshape the cellular translation landscapes. During translation elongation, ribosomes often assemble in defined three-dimensional arrangements to form polysomes4. By mapping the intracellular organization of translating ribosomes, we show that their association into polysomes involves a local coordination mechanism that is mediated by the ribosomal protein L9. We propose that an extended conformation of L9 within polysomes mitigates collisions to facilitate translation fidelity. Our work thus demonstrates the feasibility of visualizing molecular processes at atomic detail inside cells.
    DOI:  https://doi.org/10.1038/s41586-022-05255-2
  4. Biochemistry. 2022 Sep 30.
    A Tale of Water Molecules in the Ribosomal Peptidyl Transferase Reaction.
    Qiang Wang, Haibin Su.
      The peptidyl transferase center (PTC) in the large subunit of the ribosome plays a critical role in protein synthesis by catalyzing the formation of peptide bonds with an astounding speed of about 15 to 20 peptide bonds per second. The ribosome coordinates the nucleophilic attack and deprotonation in the rate-limiting step at the PTC. However, the details of peptide bond formation within the ribosome, particularly the precise role of the two water molecules in the PTC, remain unclear. Here, we propose a novel stepwise "proton shuttle" mechanism which corroborates all the reported experimental measurements so far. In this mechanism, a water molecule close to A76 of peptidyl-tRNA 2'- and 3'-O stabilizes the transition state. The other one adjacent to the carbonyl oxygen of peptidyl-tRNA actively participates in the proton shuttle, playing the catalytic role of ribosome-catalyzed peptide bond formation.
    DOI:  https://doi.org/10.1021/acs.biochem.2c00098
  5. FEBS Lett. 2022 Sep 30.
    On the role of ubiquinone in the proton translocation mechanism of respiratory complex I.
    Mårten Wikström, Amina Djurabekova, Vivek Sharma.
      Complex I converts oxidoreduction energy into a proton electrochemical gradient across the inner mitochondrial or bacterial cell membrane. This gradient is the primary source of energy for aerobic synthesis of ATP. Oxidation of reduced nicotinamide adenine dinucleotide (NADH) by ubiquinone (Q) yields NAD+ and ubiquinol (QH2 ), which is tightly coupled to translocation of four protons from the negatively to the positively charged side of the membrane. Electrons from NADH oxidation reach the iron-sulfur centre N2 positioned near the bottom of a tunnel that extends ca. 30Å from the membrane domain into the hydrophilic domain of the complex. The tunnel is occupied by ubiquinone, which can take a distal position near the N2 centre, or proximal positions closer to the membrane. Here, we review important structural, kinetic and thermodynamic properties of ubiquinone that define its role in complex I function. We suggest that this function exceeds that of a mere substrate or electron acceptor, and propose that ubiquinone may be the redox element of complex I coupling electron transfer to proton translocation.
    Keywords:  energy conservation; mitochondria; oxidative phosphorylation; proton pumping
    DOI:  https://doi.org/10.1002/1873-3468.14506
  6. Cell Rep. 2022 Sep 27. pii: S2211-1247(22)01249-9. [Epub ahead of print]40(13): 111408
    Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3.
    Ryan R Cupo, Alexandrea N Rizo, Gabriel A Braun, Eric Tse, Edward Chuang, Kushol Gupta, Daniel R Southworth, James Shorter.
      The AAA+ protein, Skd3 (human CLPB), solubilizes proteins in the mitochondrial intermembrane space, which is critical for human health. Skd3 variants with defective protein-disaggregase activity cause severe congenital neutropenia (SCN) and 3-methylglutaconic aciduria type 7 (MGCA7). How Skd3 disaggregates proteins remains poorly understood. Here, we report a high-resolution structure of a Skd3-substrate complex. Skd3 adopts a spiral hexameric arrangement that engages substrate via pore-loop interactions in the nucleotide-binding domain (NBD). Substrate-bound Skd3 hexamers stack head-to-head via unique, adaptable ankyrin-repeat domain (ANK)-mediated interactions to form dodecamers. Deleting the ANK linker region reduces dodecamerization and disaggregase activity. We elucidate apomorphic features of the Skd3 NBD and C-terminal domain that regulate disaggregase activity. We also define how Skd3 subunits collaborate to disaggregate proteins. Importantly, SCN-linked subunits sharply inhibit disaggregase activity, whereas MGCA7-linked subunits do not. These advances illuminate Skd3 structure and mechanism, explain SCN and MGCA7 inheritance patterns, and suggest therapeutic strategies.
    Keywords:  AAA+ proteins, protein aggregation; CP: Cell biology; chaperone; disaggregase; mitochondria; mitochondrial disorders; therapeutics
    DOI:  https://doi.org/10.1016/j.celrep.2022.111408
  7. STAR Protoc. 2022 Sep 30. pii: S2666-1667(22)00619-0. [Epub ahead of print]3(4): 101739
    Mass spectrometry analysis of affinity-purified cytoplasmic translation initiation complexes from human and fly cells.
    Chingakham Ranjit Singh, Naoki Tani, Akira Nakamura, Katsura Asano.
      eIF5-mimic protein (5MP) controls translation through binding to the ribosomal pre-initiation complex (PIC) and alters non-AUG translation rates for cancer oncogenes and repeat-expansions in neurodegenerative diseases. Here, we describe a semi-quantitative protocol for detecting 5MP-associated proteins in cultured human and fly cells. We detail one-step anti-FLAG affinity purification and whole-lane mass spectrometry analysis of samples resolved by SDS-PAGE. This protocol allows for quantitative evaluation of the effect of 5MP mutations on its molecular interactions, to elucidate translational control by 5MP. For complete details on the use and execution of this protocol, please refer to Singh et al. (2021).
    Keywords:  Bioinformatics; Biotechnology and bioengineering; Cell culture; Mass spectrometry; Protein biochemistry; Protein expression and purification
    DOI:  https://doi.org/10.1016/j.xpro.2022.101739
  8. Nat Rev Genet. 2022 Sep 27.
    Promoting a new view of mitochondrial genome regulation.
    Dorothy Clyde.
      
    DOI:  https://doi.org/10.1038/s41576-022-00536-y
  9. Saudi Pharm J. 2022 Aug;30(8): 1065-1078
    Recent advances in mitochondrial diseases: From molecular insights to therapeutic perspectives.
    Ahmad M Aldossary, Essam A Tawfik, Mohammed N Alomary, Samar A Alsudir, Ahmed J Alfahad, Abdullah A Alshehri, Fahad A Almughem, Rean Y Mohammed, Mai M Alzaydi.
      Mitochondria are double-membraned cytoplasmic organelles that are responsible for the production of energy in eukaryotic cells. The process is completed through oxidative phosphorylation (OXPHOS) by the respiratory chain (RC) in mitochondria. Thousands of mitochondria may be present in each cell, depending on the function of that cell. Primary mitochondria disorder (PMD) is a clinically heterogeneous disease associated with germline mutations in mitochondrial DNA (mtDNA) and/or nuclear DNA (nDNA) genes, and impairs mitochondrial structure and function. Mitochondrial dysfunction can be detected in early childhood and may be severe, progressive and often multi-systemic, involving a wide range of organs. Understanding epigenetic factors and pathways mutations can help pave the way for developing an effective cure. However, the lack of information about the disease (including age of onset, symptoms, clinical phenotype, morbidity and mortality), the limits of current preclinical models and the wide range of phenotypic presentations hamper the development of effective medicines. Although new therapeutic approaches have been introduced with encouraging preclinical and clinical outcomes, there is no definitive cure for PMD. This review highlights recent advances, particularly in children, in terms of etiology, pathophysiology, clinical diagnosis, molecular pathways and epigenetic alterations. Current therapeutic approaches, future advances and proposed new therapeutic plans will also be discussed.
    Keywords:  Epigenetic alterations; Fetal gene therapy; Gene editing; Mitochondrial diseases; Nanomedicine
    DOI:  https://doi.org/10.1016/j.jsps.2022.05.011
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