Structure of the human 80S ribosome Khatter, Heena; Myasnikov, Alexander G; Natchiar, S Kundhavai ...
Nature (London),
04/2015, Letnik:
520, Številka:
7549
Journal Article
Recenzirano
Ribosomes are translational machineries that catalyse protein synthesis. Ribosome structures from various species are known at the atomic level, but obtaining the structure of the human ribosome has ...remained a challenge; efforts to address this would be highly relevant with regard to human diseases. Here we report the near-atomic structure of the human ribosome derived from high-resolution single-particle cryo-electron microscopy and atomic model building. The structure has an average resolution of 3.6 Å, reaching 2.9 Å resolution in the most stable regions. It provides unprecedented insights into ribosomal RNA entities and amino acid side chains, notably of the transfer RNA binding sites and specific molecular interactions with the exit site tRNA. It reveals atomic details of the subunit interface, which is seen to remodel strongly upon rotational movements of the ribosomal subunits. Furthermore, the structure paves the way for analysing antibiotic side effects and diseases associated with deregulated protein synthesis.
Regulation of translation initiation is well appropriate to adapt cell growth in response to stress and environmental changes. Many bacterial mRNAs adopt structures in their 5' untranslated regions ...that modulate the accessibility of the 30S ribosomal subunit. Structured mRNAs interact with the 30S in a two-step process where the docking of a folded mRNA precedes an accommodation step. Here, we used a combination of experimental approaches in vitro (kinetic of mRNA unfolding and binding experiments to analyze mRNA-protein or mRNA-ribosome complexes, toeprinting assays to follow the formation of ribosomal initiation complexes) and in vivo (genetic) to monitor the action of ribosomal protein S1 on the initiation of structured and regulated mRNAs. We demonstrate that r-protein S1 endows the 30S with an RNA chaperone activity that is essential for the docking and the unfolding of structured mRNAs, and for the correct positioning of the initiation codon inside the decoding channel. The first three OB-fold domains of S1 retain all its activities (mRNA and 30S binding, RNA melting activity) on the 30S subunit. S1 is not required for all mRNAs and acts differently on mRNAs according to the signals present at their 5' ends. This work shows that S1 confers to the ribosome dynamic properties to initiate translation of a large set of mRNAs with diverse structural features.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Recent advances in the field of electron cryomicroscopy (cryo‐EM) have resulted in a rapidly increasing number of atomic models of biomacromolecules that have been solved using this technique and ...deposited in the Protein Data Bank and the Electron Microscopy Data Bank. Similar to macromolecular crystallography, validation tools for these models and maps are required. While some of these validation tools may be borrowed from crystallography, new methods specifically designed for cryo‐EM validation are required. Here, new computational methods and tools implemented in PHENIX are discussed, including d99 to estimate resolution, phenix.auto_sharpen to improve maps and phenix.mtriage to analyze cryo‐EM maps. It is suggested that cryo‐EM half‐maps and masks should be deposited to facilitate the evaluation and validation of cryo‐EM‐derived atomic models and maps. The application of these tools to deposited cryo‐EM atomic models and maps is also presented.
New methods and PHENIX tools for quality assessment of cryo‐EM maps, atomic models and model‐to‐map fitting are presented. Results of systematic application of these tools to high‐resolution cryo‐EM maps and corresponding atomic models are analyzed and discussed.
In structural biology, deriving and refining atomic models into maps obtained from X‐ray crystallography or cryo electron microscopy (cryo‐EM) is essential for the detailed interpretation of a ...structure and its functional implications through interactions so that for example hydrogen bonds, drug specificity and associated molecular mechanisms can be analysed. This commentary summarizes the latest features of the Phenix software and also highlights the fact that cryo‐EM increasingly contributes to data depositions in the PDB and EMDB.
In structural biology, deriving and refining atomic models into maps obtained from X‐ray crystallography or cryo electron microscopy (cryo‐EM) is essential for the detailed interpretation of a structure and its functional implications through interactions so that for example hydrogen bonds, drug specificity and associated molecular mechanisms can be analysed. This commentary summarizes the latest features of the Phenix software and also highlights the fact that cryo‐EM increasingly contributes to data depositions in the PDB and EMDB.
Abstract
The MODOMICS database was updated with recent data and now includes new data types related to RNA modifications. Changes to the database include an expanded modification catalog, ...encompassing both natural and synthetic residues identified in RNA structures. This addition aids in representing RNA sequences from the RCSB PDB database more effectively. To manage the increased number of modifications, adjustments to the nomenclature system were made. Updates in the RNA sequences section include the addition of new sequences and the reintroduction of sequence alignments for tRNAs and rRNAs. The protein section was updated and connected to structures from the RCSB PDB database and predictions by AlphaFold. MODOMICS now includes a data annotation system, with ‘Evidence’ and ‘Estimated Reliability’ features, offering clarity on data support and accuracy. This system is open to all MODOMICS entries, enhancing the accuracy of RNA modification data representation. MODOMICS is available at https://iimcb.genesilico.pl/modomics/.
Graphical Abstract
Graphical Abstract
Super-resolution microscopy (PALM, STORM etc.) provides a plethora of fluorescent signals in dense cellular environments which can be difficult to interpret. Here we describe ClusterViSu, a method ...for image reconstruction, visualization and quantification of labelled protein clusters, based on Voronoi tessellation of the individual fluorescence events. The general applicability of this clustering approach for the segmentation of super-resolution microscopy data, including for co-localization, is illustrated on a series of important biological objects such as chromatin complexes, RNA polymerase, nuclear pore complexes and microtubules.
Transcription regulation through chromatin compaction and decompaction is regulated through various chromatin-remodeling complexes such as nucleosome remodeling and histone deacetylation (NuRD) ...complex. NuRD is a 1 MDa multi-subunit protein complex which comprises many different subunits, among which histone deacetylases HDAC1/2, ATP-dependent remodeling enzymes CHD3/4, histone chaperones RbAp46/48, CpG-binding proteins MBD2/3, the GATAD2a (p66α) and/or GATAD2b (p66β) and specific DNA-binding proteins MTA1/2/3. Here, we review the currently known crystal and NMR structures of these subunits, the functional data and their relevance for biomedical research considering the implication of NuRD subunits in cancer and various other diseases. The complexity of this macromolecular assembly, and its poorly understood mode of interaction with the nucleosome, the repeating unit of chromatin, illustrate that this complex is a major challenge for structure–function relationship studies which will be tackled best by an integrated biology approach.
Chemical modifications of human ribosomal RNA (rRNA) are introduced during biogenesis and have been implicated in the dysregulation of protein synthesis, as is found in cancer and other diseases. ...However, their role in this phenomenon is unknown. Here we visualize more than 130 individual rRNA modifications in the three-dimensional structure of the human ribosome, explaining their structural and functional roles. In addition to a small number of universally conserved sites, we identify many eukaryote- or human-specific modifications and unique sites that form an extended shell in comparison to bacterial ribosomes, and which stabilize the RNA. Several of the modifications are associated with the binding sites of three ribosome-targeting antibiotics, or are associated with degenerate states in cancer, such as keto alkylations on nucleotide bases reminiscent of specialized ribosomes. This high-resolution structure of the human 80S ribosome paves the way towards understanding the role of epigenetic rRNA modifications in human diseases and suggests new possibilities for designing selective inhibitors and therapeutic drugs.
“Would it be possible to analyze molecular mechanisms and structural organisation of polyribosome assemblies using cryo electron tomography?” – we asked through a longstanding collaboration between ...my research group and that of Alexander S. Spirin. Indeed, it was: we found that double-row polyribosomes can have both circular and linear arrangements of their mRNA Afonina, Z. A., et al. (2013)
Biochemistry (Moscow)
, we figured out how eukaryotic ribosomes assemble on an mRNA to form supramolecular left-handed helices Myasnikov, A. G., et al. (2014)
Nat. Commun
., that the circularization of polyribosomes is poly-A and cap-independent Afonina, Z. A., et al. (2014)
Nucleic Acids Res.
, and that intermediary polyribosomes with open structures exist after a transition from a juvenile phase to strongly translating polysomes of medium size Afonina, Z. A., et al. (2015)
Nucleic Acids Res.
until they form densely packed helical structures with reduced activity. Our joint fruitful exchanges, hence, led to major advances in the field, which are reviewed here from a personal and historical perspective in memory of Alexander S. Spirin.
Many antibiotics in clinical use target the bacterial ribosome by interfering with the protein synthesis machinery. However, targeting the human ribosome in the case of protein synthesis ...deregulations such as in highly proliferating cancer cells has not been investigated at the molecular level up to now. Here we report the structure of the human 80S ribosome with a eukaryote-specific antibiotic and show its anti-proliferative effect on several cancer cell lines. The structure provides insights into the detailed interactions in a ligand-binding pocket of the human ribosome that are required for structure-assisted drug design. Furthermore, anti-proliferative dose response in leukaemic cells and interference with synthesis of c-myc and mcl-1 short-lived protein markers reveals specificity of a series of eukaryote-specific antibiotics towards cytosolic rather than mitochondrial ribosomes, uncovering the human ribosome as a promising cancer target.
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