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Proceedings of the National Academy of... Mar 2022SignificanceThe presence of RNA chemical modifications has long been known, but their precise molecular consequences remain unknown. 2'--methylation is an abundant...
SignificanceThe presence of RNA chemical modifications has long been known, but their precise molecular consequences remain unknown. 2'--methylation is an abundant modification that exists in RNA in all domains of life. Ribosomal RNA (rRNA) represents a functionally important RNA that is heavily modified by 2'--methylations. Although abundant at functionally important regions of the rRNA, the contribution of 2'--methylations to ribosome activities is unknown. By establishing a method to disturb rRNA 2'--methylation patterns, we show that rRNA 2'--methylations affect the function and fidelity of the ribosome and change the balance between different ribosome conformational states. Our work links 2'--methylation to ribosome dynamics and defines a set of critical rRNA 2'--methylations required for ribosome biogenesis and others that are dispensable.
Topics: Methylation; RNA; RNA, Ribosomal; Ribosomes
PubMed: 35294285
DOI: 10.1073/pnas.2117334119 -
Nature Structural & Molecular Biology Dec 2022Translation modulates the timing and amplification of gene expression after transcription. Brain development requires uniquely complex gene expression patterns, but...
Translation modulates the timing and amplification of gene expression after transcription. Brain development requires uniquely complex gene expression patterns, but large-scale measurements of translation directly in the prenatal brain are lacking. We measure the reactants, synthesis and products of mRNA translation spanning mouse neocortex neurogenesis, and discover a transient window of dynamic regulation at mid-gestation. Timed translation upregulation of chromatin-binding proteins like Satb2, which is essential for neuronal subtype differentiation, restricts protein expression in neuronal lineages despite broad transcriptional priming in progenitors. In contrast, translation downregulation of ribosomal proteins sharply decreases ribosome biogenesis, coinciding with a major shift in protein synthesis dynamics at mid-gestation. Changing activity of eIF4EBP1, a direct inhibitor of ribosome biogenesis, is concurrent with ribosome downregulation and affects neurogenesis of the Satb2 lineage. Thus, the molecular logic of brain development includes the refinement of transcriptional programs by translation. Modeling of the developmental neocortex translatome is provided as an open-source searchable resource at https://shiny.mdc-berlin.de/cortexomics .
Topics: Mice; Animals; Protein Biosynthesis; Ribosomes; Ribosomal Proteins; Codon; Brain
PubMed: 36482253
DOI: 10.1038/s41594-022-00882-9 -
Current Opinion in Chemical Biology Dec 2008Ribosome synthesis is a tightly regulated process that is crucial for cell survival. Chemical footprinting, mass spectrometry, and cryo-electron microscopy are revealing... (Review)
Review
Ribosome synthesis is a tightly regulated process that is crucial for cell survival. Chemical footprinting, mass spectrometry, and cryo-electron microscopy are revealing how these complex cellular machines are assembled. Rapid folding of the rRNA provides a platform for protein-induced assembly of the bacterial 30S ribosome. Multiple assembly pathways increase the flexibility of the assembly process, while accessory factors and modification enzymes chaperone the late stages of assembly and control the quality of the mature subunits.
Topics: Enzymes; Proteins; RNA; RNA, Ribosomal, 16S; Ribosome Subunits, Small, Bacterial; Ribosomes
PubMed: 18935976
DOI: 10.1016/j.cbpa.2008.09.024 -
International Journal of Biological... 2022Protein homeostasis is well accepted as the prerequisite for proper operation of various life activities. As the main apparatus of protein translation, ribosomes play an... (Review)
Review
Protein homeostasis is well accepted as the prerequisite for proper operation of various life activities. As the main apparatus of protein translation, ribosomes play an indispensable role in the maintenance of protein homeostasis. Nevertheless, upon stimulation of various internal and external factors, malfunction of ribosomes may be evident with the excessive production of aberrant proteins, accumulation of which can result in deleterious effects on cellular fate and even cell death. Ribosomopathies are characterized as a series of diseases caused by abnormalities of ribosomal compositions and functions. Correspondingly, cell evolves several ribosome quality control mechanisms in maintaining the quantity and quality of intracellular ribosomes, namely ribosome quality control system (RQCS). Of note, RQCS can tightly monitor the entire process from ribosome biogenesis to its degradation, with the capacity of coping with ribosomal dysfunction, including misassembled ribosomes and incorrectly synthesized ribosomal proteins. In the current literature review, we mainly introduce the RQCS and elaborate on the underlying pathogenesis of several ribosomopathies. With the in-depth understanding of ribosomal dysfunction and molecular basis of RQCS, therapeutic strategy by specifically targeting RQCS remains a promising option in treating patients with ribosomopathies and other ribosome-associated human diseases.
Topics: Eukaryota; Humans; Protein Biosynthesis; Ribosomal Proteins; Ribosomes
PubMed: 35414791
DOI: 10.7150/ijbs.70955 -
Proteomics Sep 2020Increasing evidence suggests that ribosomes actively regulate protein synthesis. However, much of this evidence is indirect, leaving this layer of gene regulation... (Review)
Review
Increasing evidence suggests that ribosomes actively regulate protein synthesis. However, much of this evidence is indirect, leaving this layer of gene regulation largely unexplored, in part due to methodological limitations. Indeed, evidence is reviewed demonstrating that commonly used methods, such as transcriptomics, are inadequate because the variability in mRNAs coding for ribosomal proteins (RP) does not necessarily correspond to RP variability. Thus protein remodeling of ribosomes should be investigated by methods that allow direct quantification of RPs, ideally of isolated ribosomes. Such methods are reviewed, focusing on mass spectrometry and emphasizing method-specific biases and approaches to control these biases. It is argued that using multiple complementary methods can help reduce the danger of interpreting reproducible systematic biases as evidence for ribosome remodeling.
Topics: Mass Spectrometry; Protein Biosynthesis; RNA, Messenger; Ribosomal Proteins; Ribosomes
PubMed: 32820594
DOI: 10.1002/pmic.202000039 -
Molecules (Basel, Switzerland) Sep 2020The extent of ribosomal heterogeneity has caught increasing interest over the past few years, as recent studies have highlighted the presence of structural variations of... (Review)
Review
The extent of ribosomal heterogeneity has caught increasing interest over the past few years, as recent studies have highlighted the presence of structural variations of the ribosome. More precisely, the heterogeneity of the ribosome covers multiple scales, including the dynamical aspects of ribosomal motion at the single particle level, specialization at the cellular and subcellular scale, or evolutionary differences across species. Upon solving the ribosome atomic structure at medium to high resolution, cryogenic electron microscopy (cryo-EM) has enabled investigating all these forms of heterogeneity. In this review, we present some recent advances in quantifying ribosome heterogeneity, with a focus on the conformational and evolutionary variations of the ribosome and their functional implications. These efforts highlight the need for new computational methods and comparative tools, to comprehensively model the continuous conformational transition pathways of the ribosome, as well as its evolution. While developing these methods presents some important challenges, it also provides an opportunity to extend our interpretation and usage of cryo-EM data, which would more generally benefit the study of molecular dynamics and evolution of proteins and other complexes.
Topics: Cell Line; Cell Membrane Permeability; Cryoelectron Microscopy; Humans; Models, Chemical; Molecular Conformation; Molecular Dynamics Simulation; Motion; Ribosomal Proteins; Ribosomes; Structure-Activity Relationship
PubMed: 32957592
DOI: 10.3390/molecules25184262 -
Current Opinion in Chemical Biology Dec 2008Protein synthesis is inherently a dynamic process, requiring both small-scale and large-scale movements of tRNA and mRNA. It has long been suspected that these movements... (Review)
Review
Protein synthesis is inherently a dynamic process, requiring both small-scale and large-scale movements of tRNA and mRNA. It has long been suspected that these movements might be coupled to conformational changes in the ribosome, and in its RNA moieties in particular. Recently, the nature of ribosome structural dynamics has begun to emerge from a combination of approaches, most notably cryo-EM, X-ray crystallography, and FRET. Ribosome movement occurs both on a grand scale, as in the intersubunit rotational movements that are coupled to tRNA-mRNA translocation, and in intricate localized rearrangements such as those that accompany codon-anticodon recognition and peptide bond formation. In spite of much progress, our understanding of the mechanics of translation is now beset with countless new questions, reflecting the vast molecular architecture of the ribosome itself.
Topics: Movement; Peptidyl Transferases; RNA, Ribosomal; Ribosome Subunits, Small; Ribosomes
PubMed: 18848900
DOI: 10.1016/j.cbpa.2008.08.037 -
Physical Review. E Jan 2022The impact of ribosome exit tunnel electrostatics on the protein elongation rate or on forces acting upon the nascent polypeptide chain are currently not fully...
The impact of ribosome exit tunnel electrostatics on the protein elongation rate or on forces acting upon the nascent polypeptide chain are currently not fully elucidated. In the past, researchers have measured the electrostatic potential inside the ribosome polypeptide exit tunnel at a limited number of spatial points, at least in rabbit reticulocytes. Here we present a basic electrostatic model of the exit tunnel of the ribosome, providing a quantitative physical description of the tunnel interaction with the nascent proteins at all centro-axial points inside the tunnel. We show that a strong electrostatic screening is due to water molecules (not mobile ions) attracted to the ribosomal nucleic acid phosphate moieties buried in the immediate vicinity of the tunnel wall. We also show how the tunnel wall components and local ribosomal protein protrusions impact on the electrostatic potential profile and impede charged amino acid residues from progressing through the tunnel, affecting the elongation rate in a range of -40% to +85% when compared to the average elongation rate. The time spent by the ribosome to decode the genetic encrypted message is constrained accordingly. We quantitatively derive, at single-residue resolution, the axial forces acting on the nascent peptide from its particular sequence embedded in the tunnel. The model sheds light on how the experimental data point measurements of the potential are linked to the local structural chemistry of the inner wall, shape, and size of the tunnel. The model consistently connects experimental observations coming from different fields in molecular biology, x-ray crystallography, physical chemistry, biomechanics, and synthetic and multiomics biology. Our model should be a valuable tool to gain insight into protein synthesis dynamics, translational control, and the role of the ribosome's mechanochemistry in the cotranslational protein folding.
Topics: Animals; Peptides; Protein Biosynthesis; Protein Folding; Rabbits; Ribosomes; Static Electricity
PubMed: 35193250
DOI: 10.1103/PhysRevE.105.014409 -
Nucleic Acids Research Sep 2022Recent studies have revealed multiple mechanisms that can lead to heterogeneity in ribosomal composition. This heterogeneity can lead to preferential translation of...
Recent studies have revealed multiple mechanisms that can lead to heterogeneity in ribosomal composition. This heterogeneity can lead to preferential translation of specific panels of mRNAs, and is defined in large part by the ribosomal protein (RP) content, amongst other things. However, it is currently unknown to what extent ribosomal composition is heterogeneous across tissues, which is compounded by a lack of tools available to study it. Here we present dripARF, a method for detecting differential RP incorporation into the ribosome using Ribosome Profiling (Ribo-seq) data. We combine the 'waste' rRNA fragment data generated in Ribo-seq with the known 3D structure of the human ribosome to predict differences in the composition of ribosomes in the material being studied. We have validated this approach using publicly available data, and have revealed a potential role for eS25/RPS25 in development. Our results indicate that ribosome heterogeneity can be detected in Ribo-seq data, providing a new method to study this phenomenon. Furthermore, with dripARF, previously published Ribo-seq data provides a wealth of new information, allowing the identification of RPs of interest in many disease and normal contexts. dripARF is available as part of the ARF R package and can be accessed through https://github.com/fallerlab/ARF.
Topics: Humans; RNA, Messenger; RNA, Ribosomal; Ribosomal Proteins; Ribosomes
PubMed: 35687114
DOI: 10.1093/nar/gkac484 -
Phytochemistry Oct 2022Ribosome inactivating proteins (RIPs) are rRNA N-glycosylases (EC 3.2.2.22) best known for hydrolyzing an adenine base from the conserved sarcin/ricin loop of ribosomal... (Review)
Review
Ribosome inactivating proteins (RIPs) are rRNA N-glycosylases (EC 3.2.2.22) best known for hydrolyzing an adenine base from the conserved sarcin/ricin loop of ribosomal RNA. Protein translation is inhibited by ribosome depurination; therefore, RIPs are generally considered toxic to cells. The expression of some RIPs is upregulated by biotic and abiotic stress, though the connection between RNA depurination and defense response is not well understood. Despite their prevalence in approximately one-third of flowering plant orders, our knowledge of RIPs stems primarily from biochemical analyses of individuals or genomics-scale analyses of small datasets from a limited number of species. Here, we performed an unbiased search for proteins with RIP domains and identified several-fold more RIPs than previously known - more than 800 from 120 species, many with novel associated domains and physicochemical characteristics. Based on protein domain configuration, we established 15 distinct groups, suggesting diverse functionality. Surprisingly, most of these RIPs lacked a signal peptide, indicating they may be localized to the nucleocytoplasm of cells, raising questions regarding their toxicity against conspecific ribosomes. Our phylogenetic analysis significantly extends previous models for RIP evolution in plants, predicting an original single-domain RIP that later evolved to acquire a signal peptide and different protein domains. We show that RIPs are distributed throughout 21 plant orders with many species maintaining genes for more than one RIP group. Our analyses provide the foundation for further characterization of these new RIP types, to understand how these enzymes function in plants.
Topics: Phylogeny; Plant Proteins; Protein Sorting Signals; RNA, Ribosomal; Ribosome Inactivating Proteins; Ribosomes
PubMed: 35934106
DOI: 10.1016/j.phytochem.2022.113337