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ELife Feb 2024A new in vitro system called Rec-Seq sheds light on how mRNA molecules compete for the machinery that translates their genetic sequence into proteins.
A new in vitro system called Rec-Seq sheds light on how mRNA molecules compete for the machinery that translates their genetic sequence into proteins.
Topics: Protein Biosynthesis; Ribosomes; RNA, Messenger
PubMed: 38393777
DOI: 10.7554/eLife.96304 -
Current Opinion in Chemical Biology Jun 2024Novel discoveries in natural product biosynthesis reveal hidden bioactive compounds and expand our knowledge in enzymology. Ribosomally synthesized and... (Review)
Review
Novel discoveries in natural product biosynthesis reveal hidden bioactive compounds and expand our knowledge in enzymology. Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a rapidly growing class of natural products featuring diverse non-canonical amino acids introduced by maturation enzymes as a class-defining characteristic. Underexplored RiPP sources, such as the human microbiome, the oceans, uncultured microorganisms, and plants are rich hunting grounds for novel enzymology. Unusual α- and β-amino acids, peptide cleavages, lipidations, diverse macrocyclizations, and other features expand the range of chemical groups that are installed in RiPPs by often promiscuous enzymes. This review highlights the search for novelty in RiPP enzymology in the past two years, with respect to the discovery of new biochemical modifications but also towards novel applications.
Topics: Protein Processing, Post-Translational; Humans; Peptides; Ribosomes; Biological Products; Animals; Enzymes
PubMed: 38729090
DOI: 10.1016/j.cbpa.2024.102463 -
Cell Reports Sep 2023Suppression of premature termination codons (PTCs) by translational readthrough is a promising strategy to treat a wide variety of severe genetic diseases caused by...
Suppression of premature termination codons (PTCs) by translational readthrough is a promising strategy to treat a wide variety of severe genetic diseases caused by nonsense mutations. Here, we present two potent readthrough promoters-NVS1.1 and NVS2.1-that restore substantial levels of functional full-length CFTR and IDUA proteins in disease models for cystic fibrosis and Hurler syndrome, respectively. In contrast to other readthrough promoters that affect stop codon decoding, the NVS compounds stimulate PTC suppression by triggering rapid proteasomal degradation of the translation termination factor eRF1. Our results show that this occurs by trapping eRF1 in the terminating ribosome, causing ribosome stalls and subsequent ribosome collisions, and activating a branch of the ribosome-associated quality control network, which involves the translational stress sensor GCN1 and the catalytic activity of the E3 ubiquitin ligases RNF14 and RNF25.
Topics: Humans; Codon, Terminator; Protein Biosynthesis; Codon, Nonsense; Ribosomes; Cystic Fibrosis
PubMed: 37651229
DOI: 10.1016/j.celrep.2023.113056 -
Experimental & Molecular Medicine Jul 2023Transfer RNAs (tRNAs) play an essential role in mRNA translation by delivering amino acids to growing polypeptide chains. Recent data demonstrate that tRNAs can be... (Review)
Review
Transfer RNAs (tRNAs) play an essential role in mRNA translation by delivering amino acids to growing polypeptide chains. Recent data demonstrate that tRNAs can be cleaved by ribonucleases, and the resultant cleavage products, tRNA-derived small RNAs (tsRNAs), have crucial roles in physiological and pathological conditions. They are classified into more than six types according to their size and cleavage positions. Since the initial discovery of the physiological functions of tsRNAs more than a decade ago, accumulating data have demonstrated that tsRNAs play critical roles in gene regulation and tumorigenesis. These tRNA-derived molecules have various regulatory functions at the transcriptional, post-transcriptional, and translational levels. More than a hundred types of modifications are found on tRNAs, affecting the biogenesis, stability, function, and biochemical properties of tsRNA. Both oncogenic and tumor suppressor functions have been reported for tsRNAs, which play important roles in the development and progression of various cancers. Abnormal expression patterns and modification of tsRNAs are associated with various diseases, including cancer and neurological disorders. In this review, we will describe the biogenesis, versatile gene regulation mechanisms, and modification-mediated regulation mechanisms of tsRNA as well as the expression patterns and potential therapeutic roles of tsRNAs in various cancers.
Topics: Humans; RNA, Transfer; Neoplasms; Protein Biosynthesis; Gene Expression Regulation; Biology
PubMed: 37430089
DOI: 10.1038/s12276-023-01038-5 -
Accounts of Chemical Research Oct 2023Protein biosynthesis is a central process in all living cells that is catalyzed by a complex molecular machine─the ribosome. This process is termed translation because... (Review)
Review
Protein biosynthesis is a central process in all living cells that is catalyzed by a complex molecular machine─the ribosome. This process is termed translation because the language of nucleotides in mRNAs is translated into the language of amino acids in proteins. Transfer RNA (tRNA) molecules charged with amino acids serve as adaptors and recognize codons of mRNA in the decoding center while simultaneously the individual amino acids are assembled into a peptide chain in the peptidyl transferase center (PTC). As the nascent peptide emerges from the ribosome, it is threaded through a long tunnel referred to as a nascent peptide exit tunnel (NPET). The PTC and NPET are the sites targeted by many antibiotics and are thus of tremendous importance from a biomedical perspective and for drug development in the pharmaceutical industry.Researchers have achieved much progress in characterizing ribosomal translation at the molecular level; an impressive number of high-resolution structures of different functional and inhibited states of the ribosome are now available. These structures have significantly contributed to our understanding of how the ribosome interacts with its key substrates, namely, mRNA, tRNAs, and translation factors. In contrast, much less is known about the mechanisms of how small molecules, especially antibiotics, affect ribosomal protein synthesis. This mainly concerns the structural basis of small molecule-NPET interference with cotranslational protein folding and the regulation of protein synthesis. Growing biochemical evidence suggests that NPET plays an active role in the regulation of protein synthesis.Much-needed progress in this field is hampered by the fact that during the preparation of ribosome complexes for structural studies (i.e., X-ray crystallography, cryoelectron microscopy, and NMR spectroscopy) the aminoacyl- or peptidyl-tRNAs are unstable and become hydrolyzed. A solution to this problem is the application of hydrolysis-resistant mimics of aminoacyl- or peptidyl-tRNAs.In this Account, we present an overview of synthetic methods for the generation of peptidyl-tRNA analogs. Modular approaches have been developed that combine () RNA and peptide solid-phase synthesis on 3'-aminoacylamino-adenosine resins, () native chemical ligations and Staudinger ligations, () tailoring of tRNAs by the selective cleavage of natural native tRNAs with DNAzymes followed by reassembly with enzymatic ligation to synthetic peptidyl-RNA fragments, and () enzymatic tailing and cysteine charging of the tRNA to obtain modified CCA termini of a tRNA that are chemically ligated to the peptide moiety of interest. With this arsenal of tools, in principle, any desired sequence of a stably linked peptidyl-tRNA mimic is accessible. To underline the significance of the synthetic conjugates, we briefly point to the most critical applications that have shed new light on the molecular mechanisms underlying the context-specific activity of ribosome-targeting antibiotics, ribosome-dependent incorporation of multiple consecutive proline residues, the incorporation of d-amino acids, and tRNA mischarging.Furthermore, we discuss new types of stably charged tRNA analogs, relying on triazole- and squarate (instead of amide)-linked conjugates. Those have pushed forward our mechanistic understanding of nonribosomal peptide synthesis, where aminoacyl-tRNA-dependent enzymes are critically involved in various cellular processes in primary and secondary metabolism and in bacterial cell wall synthesis.
Topics: Cryoelectron Microscopy; RNA, Transfer; RNA; Amino Acids; Protein Biosynthesis; Peptides; Anti-Bacterial Agents; RNA, Messenger; Biology
PubMed: 37728742
DOI: 10.1021/acs.accounts.3c00412 -
Journal of Experimental Botany Oct 2023In response to changing environmental conditions, plants activate cellular responses to enable them to adapt. One such response is autophagy, in which cellular... (Review)
Review
In response to changing environmental conditions, plants activate cellular responses to enable them to adapt. One such response is autophagy, in which cellular components, for example proteins and organelles, are delivered to the vacuole for degradation. Autophagy is activated by a wide range of conditions, and the regulatory pathways controlling this activation are now being elucidated. However, key aspects of how these factors may function together to properly modulate autophagy in response to specific internal or external signals are yet to be discovered. In this review we discuss mechanisms for regulation of autophagy in response to environmental stress and disruptions in cell homeostasis. These pathways include post-translational modification of proteins required for autophagy activation and progression, control of protein stability of the autophagy machinery, and transcriptional regulation, resulting in changes in transcription of genes involved in autophagy. In particular, we highlight potential connections between the roles of key regulators and explore gaps in research, the filling of which can further our understanding of the autophagy regulatory network in plants.
Topics: Autophagy; Gene Expression Regulation; Protein Processing, Post-Translational; Homeostasis; Vacuoles
PubMed: 37358252
DOI: 10.1093/jxb/erad211 -
International Journal of Molecular... Feb 2024In recent years, research into Parkinson's disease and similar neurodegenerative disorders has increasingly suggested that these conditions are synonymous with failures... (Review)
Review
In recent years, research into Parkinson's disease and similar neurodegenerative disorders has increasingly suggested that these conditions are synonymous with failures in proteostasis. However, the spotlight of this research has remained firmly focused on the tail end of proteostasis, primarily aggregation, misfolding, and degradation, with protein translation being comparatively overlooked. Now, there is an increasing body of evidence supporting a potential role for translation in the pathogenesis of PD, and its dysregulation is already established in other similar neurodegenerative conditions. In this paper, we consider how altered protein translation fits into the broader picture of PD pathogenesis, working hand in hand to compound the stress placed on neurons, until this becomes irrecoverable. We will also consider molecular players of interest, recent evidence that suggests that aggregates may directly influence translation in PD progression, and the implications for the role of protein translation in our development of clinically useful diagnostics and therapeutics.
Topics: Humans; Parkinson Disease; Neurons; Proteostasis; Protein Biosynthesis; alpha-Synuclein
PubMed: 38397070
DOI: 10.3390/ijms25042393 -
Aging Cell Sep 2023Cellular senescence constitutes a generally irreversible proliferation barrier, accompanied by macromolecular damage and metabolic rewiring. Several senescence types...
Cellular senescence constitutes a generally irreversible proliferation barrier, accompanied by macromolecular damage and metabolic rewiring. Several senescence types have been identified based on the initiating stimulus, such as replicative (RS), stress-induced (SIS) and oncogene-induced senescence (OIS). These senescence subtypes are heterogeneous and often develop subset-specific phenotypes. Reduced protein synthesis is considered a senescence hallmark, but whether this trait pertains to various senescence subtypes and if distinct molecular mechanisms are involved remain largely unknown. Here, we analyze large published or experimentally produced RNA-seq and Ribo-seq datasets to determine whether major translation-regulating entities such as ribosome stalling, the presence of uORFs/dORFs and IRES elements may differentially contribute to translation deficiency in senescence subsets. We show that translation-regulating mechanisms may not be directly relevant to RS, however uORFs are significantly enriched in SIS. Interestingly, ribosome stalling, uORF/dORF patterns and IRES elements comprise predominant mechanisms upon OIS, strongly correlating with Notch pathway activation. Our study provides for the first time evidence that major translation dysregulation mechanisms/patterns occur during cellular senescence, but at different rates depending on the stimulus type. The degree at which those mechanisms accumulate directly correlates with translation deficiency levels. Our thorough analysis contributes to elucidating crucial and so far unknown differences in the translation machinery between senescence subsets.
Topics: Cellular Senescence; Ribosomes; Protein Biosynthesis
PubMed: 37547972
DOI: 10.1111/acel.13893 -
Proceedings of the National Academy of... Dec 2023Activation of neuronal protein synthesis upon learning is critical for the formation of long-term memory. Here, we report that learning in the contextual fear...
Activation of neuronal protein synthesis upon learning is critical for the formation of long-term memory. Here, we report that learning in the contextual fear conditioning paradigm engenders a decrease in eIF2α (eukaryotic translation initiation factor 2) phosphorylation in astrocytes in the hippocampal CA1 region, which promotes protein synthesis. Genetic reduction of eIF2α phosphorylation in hippocampal astrocytes enhanced contextual and spatial memory and lowered the threshold for the induction of long-lasting plasticity by modulating synaptic transmission. Thus, learning-induced dephosphorylation of eIF2α in astrocytes bolsters hippocampal synaptic plasticity and consolidation of long-term memories.
Topics: Long-Term Potentiation; Astrocytes; Neuronal Plasticity; Hippocampus; Protein Biosynthesis; CA1 Region, Hippocampal; Memory, Long-Term
PubMed: 38015848
DOI: 10.1073/pnas.2308671120 -
The Journal of Biological Chemistry Sep 2023Iron-sulfur clusters (ISC) are essential cofactors that participate in electron transfer, environmental sensing, and catalysis. Amongst the most ancient ISC-containing...
Iron-sulfur clusters (ISC) are essential cofactors that participate in electron transfer, environmental sensing, and catalysis. Amongst the most ancient ISC-containing proteins are the ferredoxin (FDX) family of electron carriers. Humans have two FDXs- FDX1 and FDX2, both of which are localized to mitochondria, and the latter of which is itself important for ISC synthesis. We have previously shown that hypoxia can eliminate the requirement for some components of the ISC biosynthetic pathway, but FDXs were not included in that study. Here, we report that FDX1, but not FDX2, is dispensable under 1% O in cultured human cells. We find that FDX1 is essential for production of the lipoic acid cofactor, which is synthesized by the ISC-containing enzyme lipoyl synthase. While hypoxia can rescue the growth phenotype of either FDX1 or lipoyl synthase KO cells, lipoylation in these same cells is not rescued, arguing against an alternative biosynthetic route or salvage pathway for lipoate in hypoxia. Our work reveals the divergent roles of FDX1 and FDX2 in mitochondria, identifies a role for FDX1 in lipoate synthesis, and suggests that loss of lipoic acid can be tolerated under low oxygen tensions in cell culture.
Topics: Humans; Ferredoxins; Lipoylation; Thioctic Acid; Cell Hypoxia; Gene Knockout Techniques; Oxygen; Proteome; Sulfurtransferases; Binding Sites; Protein Stability; Protein Biosynthesis
PubMed: 37481209
DOI: 10.1016/j.jbc.2023.105075