Search

Ribosomal RNAs (rRNAs) are extensively modified during the transcription and subsequent maturation. Three types of modifications, 2′-O-methylation of ribose moiety, pseudouridylation, and base modifications, are introduced either by a snoRNA-driven mechanism or by stand-alone enzymes. Modified nucleotides are clustered at the functionally important sites, including peptidyl transferase center (PTC). Therefore, it has been hypothesised that the modified nucleotides play an important role in ensuring the functionality of the ribosome. In this study, we demonstrate that seven 25S rRNA modifications, including four evolutionarily conserved modifications, in the proximity of PTC can be simultaneously depleted without loss of cell viability. Yeast mutants lacking three snoRNA genes (snR34, snR52, and snR65) and/or expressing enzymatically inactive variants of spb1(D52A/E679K) and nop2(C424A/C478A) were constructed. The results show that rRNA modifications in PTC contribute collectively to efficient translation in eukaryotic cells. The deficiency of seven modified nucleotides in 25S rRNA resulted in reduced cell growth, cold sensitivity, decreased translation levels, and hyperaccurate translation, as indicated by the reduced missense and nonsense suppression. The modification m5C2870 is crucial in the absence of the other six modified nucleotides. Thus, the pattern of rRNA-modified nucleotides around the PTC is essential for optimal ribosomal translational activity and translational fidelity. [ABSTRACT FROM AUTHOR]

For over 40 years, ribosomes were considered monolithic machines that translate the genetic code indiscriminately. However, over the past two decades, there have been a growing number of studies that suggest ribosomes to have a degree of compositional and functional adaptability in response to tissue type, cell environment and stimuli, cell cycle or development state. In such form, ribosomes themselves take an active part in translation regulation through an intrinsic adaptability provided by evolution, which furnished ribosomes with a dynamic plasticity that confers another layer of gene expression regulation. Yet despite the identification of various sources that give rise to ribosomal heterogeneity both at the protein and RNA level, its functional relevance is still debated, and many questions remain. Here, we will review aspects, including evolutionary ones, of ribosome heterogeneity emerging at the nucleic acid level, and aim to reframe ribosome ‘heterogeneity’ as an adaptive and dynamic process of plasticity.The terms on which this article has been published allow the posting of the Accepted Manuscript in a repository by the author(s) or with their consent.

Since the discovery of the first ribonucleic acid (RNA) modifications in transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs), scientists have been on a quest to decipher the identities and functions of RNA modifications in biological systems. The last decade has seen monumental growth in the number of studies that have characterized and assessed the functionalities of RNA modifications in the field of plant biology. Owing to these studies, we now categorize RNA modifications based on their chemical nature and the RNA on which they are found, as well as the array of proteins that are involved in the processes that add, read, and remove them from an RNA molecule. Beyond their identity, another key piece of the puzzle is the functional significance of the various types of RNA modifications. Here, we shed light on recent studies that help establish our current understanding of the diversity of RNA modifications found in plant transcriptomes and the functions they play at both the molecular (e.g., RNA stability, translation, and transport) and organismal (e.g., stress response and development) levels. Finally, we consider the key research questions related to plant gene expression and biology in general and highlight developments in various technologies that are driving our insights forward in this research area. [ABSTRACT FROM AUTHOR]

For over 40 years, ribosomes were considered monolithic machines that translate the genetic code indiscriminately. However, over the past two decades, there have been a growing number of studies that suggest ribosomes to have a degree of compositional and functional adaptability in response to tissue type, cell environment and stimuli, cell cycle or development state. In such form, ribosomes themselves take an active part in translation regulation through an intrinsic adaptability provided by evolution, which furnished ribosomes with a dynamic plasticity that confers another layer of gene expression regulation. Yet despite the identification of various sources that give rise to ribosomal heterogeneity both at the protein and RNA level, its functional relevance is still debated, and many questions remain. Here, we will review aspects, including evolutionary ones, of ribosome heterogeneity emerging at the nucleic acid level, and aim to reframe ribosome 'heterogeneity' as an adaptive and dynamic process of plasticity.The terms on which this article has been published allow the posting of the Accepted Manuscript in a repository by the author(s) or with their consent. [ABSTRACT FROM AUTHOR]

The conceptual origins of ribosome specialization can be traced back to the earliest days of molecular biology. Yet, this field has only recently begun to gather momentum, with numerous studies identifying distinct heterogeneous ribosome populations across multiple species and model systems. It is proposed that some of these compositionally distinct ribosomes may be functionally specialized and able to regulate the translation of specific mRNAs. Identification and functional characterization of specialized ribosomes has the potential to elucidate a novel layer of gene expression control, at the level of translation, where the ribosome itself is a key regulatory player. In this review, we discuss different sources of ribosome heterogeneity, evidence for ribosome specialization, and also the future directions of this exciting field. Over the past decade, it has become increasingly apparent that ribosomes are significantly more heterogeneous than originally thought, with variation emanating from both the rRNA and protein content of ribosomes. This heterogeneity can potentially confer functional ribosome specialization, contributing to translational control, thereby identifying the ribosome as a key regulatory player in translation. Studies have uncovered interindividual and intertissue variant rRNA alleles and there has also been a recent focus on identifying functional roles for the previously enigmatic eukaryotic rRNA expansion segments. Technological advances have permitted accurate, quantitative analysis of rRNA modifications across several organisms, with functionality ascribed to certain rRNA modifications. Other work has centered on identifying specialization arising due to changes in ribosome protein composition. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a bacterial pathogen and one of the leading causes of healthcare‐acquired infections in the world. The growing antibiotic resistance of S. aureus obliges us to search for new drugs and treatments. As the majority of antibiotics target the ribosome, knowledge of its detailed structure is crucial for drug development. Here, we report the cryo‐EM reconstruction at 3.2 Å resolution of the S. aureus ribosome with P‐site tRNA, messenger RNA, and 10 RNA modification sites previously not assigned or visualized. The resulting model is the most precise and complete high‐resolution structure to date of the S. aureus 70S ribosome with functional ligands. [ABSTRACT FROM AUTHOR]

Background: Antibiotic resistance is a growing worldwide problem. One of the major resistant bacterial pathogens is Staphylococcus aureus, which became a burden of healthcare systems around the world. To overcome the issue, more drug discovery studies are needed. One of the main antibiotic targets is a ribosome – the central hub of protein synthesis. Structural data of the ribosome and its features are a crucial milestone for the effective development of new drugs, especially using structure-based drug design approaches. Apart from many small structural features, ribosome possesses rRNA modifications that play a role in the fine-tuning of protein synthesis. Detailed species-specific structural data of the S. aureus ribosome is also a useful model for understanding the resistance mechanisms. This information could help with the design of new antibiotics and the upgrading of old ones. The data on S. aureus ribosomal RNA modifications and corresponding modification enzymes are very limited. Our aim was to improve the current models of the S. aureus ribosome by determining its structure with functional ligands at a much higher resolution - thereby creating a foundation for structure-based drug design experiments and research of new drug targets. Methods: The S. aureus ribosome complex consists of three components: ribosome, fMet-tRNAfMet, mRNA and 70S ribosome. The complex from purified components was formed in vitro and applied to cryo-EM grids. Data was collected at Titan Krios with Gatan K2 detector (IGBMC, France). The data was processed and modeled in Relion 2.1, Chimera, Coot, and Phenix. Results: We determined the cryo-EM reconstruction at 3.2 Å resolution of the S. aureus ribosome with P-site tRNA, messenger RNA. Based on the experimental map and existing bioinformatic data, we at the first time identified and assigned 10 modifications of S. aureus rRNA. We analyzed the positions of rRNA modifications and their possible functions. Conclusion: In this study, we describe our structure of S. aureus ribosome with functional ligands. The present model is the highest resolution and most precise that is available at the moment. We propose a set of methyltransferases as targets for future drug discovery studies. The proposed methyltransferases and corresponding modifications may play an important role in protein synthesis and its regulation.

Embryonic stem cells (ESCs) and adult stem cells (ASCs) possess the remarkable capacity to self-renew while remaining poised to differentiate into multiple progenies in the context of a rapidly developing embryo or in steady-state tissues, respectively. This ability is controlled by complex genetic programs, which are dynamically orchestrated at different steps of gene expression, including chromatin remodeling, mRNA transcription, processing, and stability. In addition to maintaining stem cell homeostasis, these molecular processes need to be rapidly rewired to coordinate complex physiological modifications required to redirect cell fate in response to environmental clues, such as differentiation signals or tissue injuries. Although chromatin remodeling and mRNA expression have been extensively studied in stem cells, accumulating evidence suggests that stem cell transcriptomes and proteomes are poorly correlated and that stem cell properties require finely tuned protein synthesis. In addition, many studies have shown that the biogenesis of the translation machinery, the ribosome, is decisive for sustaining ESC and ASC properties. Therefore, these observations emphasize the importance of translational control in stem cell homeostasis and fate decisions. In this review, we will provide the most recent literature describing how ribosome biogenesis and translational control regulate stem cell functions and are crucial for accommodating proteome remodeling in response to changes in stem cell fate.

In human ribosomal RNAs, over 200 residues are modified by specific, RNA-driven enzymatic complexes or stand-alone, RNA-independent enzymes. In most cases, modification sites are placed in specific positions within important functional areas of the ribosome. Some evidence indicates that the altered control in ribosomal RNA modifications may affect ribosomal function during mRNA translation. Here we provide an overview of the connections linking ribosomal RNA modifications to ribosome function, and suggest how aberrant modifications may affect the control of the expression of key cancer genes, thus contributing to tumor development. In addition, the future perspectives in this field are discussed. [ABSTRACT FROM AUTHOR]

The ability to detect 2'-O-methylation sites (Nm) in high-throughput fashion is important, as increasing evidence points to a more diverse landscape for this RNA modification as well as the possibility of yet unidentified functions. Here we describe an optimized version of RibOxi-seq, which is built upon the original published method, that not only accurately profiles ribosomal RNA (rRNA) Nm sites with minimal RNA input but is also robust enough to identify mRNA intronic and exonic sites., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Embryonic stem cells (ESCs) and adult stem cells (ASCs) possess the remarkable capacity to self-renew while remaining poised to differentiate into multiple progenies in the context of a rapidly developing embryo or in steady-state tissues, respectively. This ability is controlled by complex genetic programs, which are dynamically orchestrated at different steps of gene expression, including chromatin remodeling, mRNA transcription, processing, and stability. In addition to maintaining stem cell homeostasis, these molecular processes need to be rapidly rewired to coordinate complex physiological modifications required to redirect cell fate in response to environmental clues, such as differentiation signals or tissue injuries. Although chromatin remodeling and mRNA expression have been extensively studied in stem cells, accumulating evidence suggests that stem cell transcriptomes and proteomes are poorly correlated and that stem cell properties require finely tuned protein synthesis. In addition, many studies have shown that the biogenesis of the translation machinery, the ribosome, is decisive for sustaining ESC and ASC properties. Therefore, these observations emphasize the importance of translational control in stem cell homeostasis and fate decisions. In this review, we will provide the most recent literature describing how ribosome biogenesis and translational control regulate stem cell functions and are crucial for accommodating proteome remodeling in response to changes in stem cell fate. [ABSTRACT FROM AUTHOR]