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Nature Reviews. Molecular Cell Biology Feb 2022In eukaryotes, poly(A) tails are present on almost every mRNA. Early experiments led to the hypothesis that poly(A) tails and the cytoplasmic polyadenylate-binding... (Review)
Review
In eukaryotes, poly(A) tails are present on almost every mRNA. Early experiments led to the hypothesis that poly(A) tails and the cytoplasmic polyadenylate-binding protein (PABPC) promote translation and prevent mRNA degradation, but the details remained unclear. More recent data suggest that the role of poly(A) tails is much more complex: poly(A)-binding protein can stimulate poly(A) tail removal (deadenylation) and the poly(A) tails of stable, highly translated mRNAs at steady state are much shorter than expected. Furthermore, the rate of translation elongation affects deadenylation. Consequently, the interplay between poly(A) tails, PABPC, translation and mRNA decay has a major role in gene regulation. In this Review, we discuss recent work that is revolutionizing our understanding of the roles of poly(A) tails in the cytoplasm. Specifically, we discuss the roles of poly(A) tails in translation and control of mRNA stability and how poly(A) tails are removed by exonucleases (deadenylases), including CCR4-NOT and PAN2-PAN3. We also discuss how deadenylation rate is determined, the integration of deadenylation with other cellular processes and the function of PABPC. We conclude with an outlook for the future of research in this field.
Topics: Animals; Eukaryota; Gene Expression Regulation; Humans; Poly A; Protein Biosynthesis; RNA Stability; RNA, Messenger
PubMed: 34594027
DOI: 10.1038/s41580-021-00417-y -
RNA (New York, N.Y.) Aug 2020The aminoacyl-tRNA synthetases are an essential and universally distributed family of enzymes that plays a critical role in protein synthesis, pairing tRNAs with their... (Review)
Review
The aminoacyl-tRNA synthetases are an essential and universally distributed family of enzymes that plays a critical role in protein synthesis, pairing tRNAs with their cognate amino acids for decoding mRNAs according to the genetic code. Synthetases help to ensure accurate translation of the genetic code by using both highly accurate cognate substrate recognition and stringent proofreading of noncognate products. While alterations in the quality control mechanisms of synthetases are generally detrimental to cellular viability, recent studies suggest that in some instances such changes facilitate adaption to stress conditions. Beyond their central role in translation, synthetases are also emerging as key players in an increasing number of other cellular processes, with far-reaching consequences in health and disease. The biochemical versatility of the synthetases has also proven pivotal in efforts to expand the genetic code, further emphasizing the wide-ranging roles of the aminoacyl-tRNA synthetase family in synthetic and natural biology.
Topics: Amino Acyl-tRNA Synthetases; Animals; Genetic Code; Humans; Protein Biosynthesis; RNA, Messenger; RNA, Transfer; Transfer RNA Aminoacylation
PubMed: 32303649
DOI: 10.1261/rna.071720.119 -
Nature Reviews. Molecular Cell Biology May 2021Mitochondria are cellular organelles responsible for generation of chemical energy in the process called oxidative phosphorylation. They originate from a bacterial... (Review)
Review
Mitochondria are cellular organelles responsible for generation of chemical energy in the process called oxidative phosphorylation. They originate from a bacterial ancestor and maintain their own genome, which is expressed by designated, mitochondrial transcription and translation machineries that differ from those operating for nuclear gene expression. In particular, the mitochondrial protein synthesis machinery is structurally and functionally very different from that governing eukaryotic, cytosolic translation. Despite harbouring their own genetic information, mitochondria are far from being independent of the rest of the cell and, conversely, cellular fitness is closely linked to mitochondrial function. Mitochondria depend heavily on the import of nuclear-encoded proteins for gene expression and function, and hence engage in extensive inter-compartmental crosstalk to regulate their proteome. This connectivity allows mitochondria to adapt to changes in cellular conditions and also mediates responses to stress and mitochondrial dysfunction. With a focus on mammals and yeast, we review fundamental insights that have been made into the biogenesis, architecture and mechanisms of the mitochondrial translation apparatus in the past years owing to the emergence of numerous near-atomic structures and a considerable amount of biochemical work. Moreover, we discuss how cellular mitochondrial protein expression is regulated, including aspects of mRNA and tRNA maturation and stability, roles of auxiliary factors, such as translation regulators, that adapt mitochondrial translation rates, and the importance of inter-compartmental crosstalk with nuclear gene expression and cytosolic translation and how it enables integration of mitochondrial translation into the cellular context.
Topics: Animals; Eukaryota; Gene Expression Regulation; Humans; Mitochondria; Oxidative Phosphorylation; Protein Biosynthesis; RNA, Messenger; RNA, Transfer; Transcription, Genetic
PubMed: 33594280
DOI: 10.1038/s41580-021-00332-2 -
British Journal of Cancer Jan 2020An abundant supply of amino acids is important for cancers to sustain their proliferative drive. Alongside their direct role as substrates for protein synthesis, they... (Review)
Review
An abundant supply of amino acids is important for cancers to sustain their proliferative drive. Alongside their direct role as substrates for protein synthesis, they can have roles in energy generation, driving the synthesis of nucleosides and maintenance of cellular redox homoeostasis. As cancer cells exist within a complex and often nutrient-poor microenvironment, they sometimes exist as part of a metabolic community, forming relationships that can be both symbiotic and parasitic. Indeed, this is particularly evident in cancers that are auxotrophic for particular amino acids. This review discusses the stromal/cancer cell relationship, by using examples to illustrate a number of different ways in which cancer cells can rely on and contribute to their microenvironment - both as a stable network and in response to therapy. In addition, it examines situations when amino acid synthesis is driven through metabolic coupling to other reactions, and synthesis is in excess of the cancer cell's proliferative demand. Finally, it highlights the understudied area of non-proteinogenic amino acids in cancer metabolism and their potential role.
Topics: Amino Acids; Cell Proliferation; Energy Metabolism; Humans; Neoplasms; Protein Biosynthesis; Tumor Microenvironment
PubMed: 31819187
DOI: 10.1038/s41416-019-0620-5 -
Proceedings of the National Academy of... Nov 2019Messenger RNAs (mRNAs) encode information in both their primary sequence and their higher order structure. The independent contributions of factors like codon usage and...
Messenger RNAs (mRNAs) encode information in both their primary sequence and their higher order structure. The independent contributions of factors like codon usage and secondary structure to regulating protein expression are difficult to establish as they are often highly correlated in endogenous sequences. Here, we used 2 approaches, global inclusion of modified nucleotides and rational sequence design of exogenously delivered constructs, to understand the role of mRNA secondary structure independent from codon usage. Unexpectedly, highly expressed mRNAs contained a highly structured coding sequence (CDS). Modified nucleotides that stabilize mRNA secondary structure enabled high expression across a wide variety of primary sequences. Using a set of eGFP mRNAs with independently altered codon usage and CDS structure, we find that the structure of the CDS regulates protein expression through changes in functional mRNA half-life (i.e., mRNA being actively translated). This work highlights an underappreciated role of mRNA secondary structure in the regulation of mRNA stability.
Topics: Half-Life; HeLa Cells; Humans; Nucleic Acid Conformation; Protein Biosynthesis; Proteins; RNA Stability; RNA, Messenger
PubMed: 31712433
DOI: 10.1073/pnas.1908052116 -
Trends in Cell Biology Jun 2020Eukaryotic cells must accurately monitor the integrity of the mitochondrial network to overcome environmental insults and respond to physiological cues. The... (Review)
Review
Eukaryotic cells must accurately monitor the integrity of the mitochondrial network to overcome environmental insults and respond to physiological cues. The mitochondrial unfolded protein response (UPR) is a mitochondrial-to-nuclear signaling pathway that maintains mitochondrial proteostasis, mediates signaling between tissues, and regulates organismal aging. Aberrant UPR signaling is associated with a wide spectrum of disorders, including congenital diseases as well as cancers and neurodegenerative diseases. Here, we review recent research into the mechanisms underlying UPR signaling in Caenorhabditis elegans and discuss emerging connections between the UPR signaling and a translational regulation program called the 'integrated stress response'. Further study of the UPR will potentially enable development of new therapeutic strategies for inherited metabolic disorders and diseases of aging.
Topics: Animals; Humans; Mitochondria; Protein Biosynthesis; Signal Transduction; Stress, Physiological; Unfolded Protein Response
PubMed: 32413314
DOI: 10.1016/j.tcb.2020.03.001 -
Circulation Nov 2021The integrated stress response (ISR) is an evolutionarily conserved process to cope with intracellular and extracellular disturbances. Myocardial infarction is a leading...
BACKGROUND
The integrated stress response (ISR) is an evolutionarily conserved process to cope with intracellular and extracellular disturbances. Myocardial infarction is a leading cause of death worldwide. Coronary artery reperfusion, the most effective means to mitigate cardiac damage of myocardial infarction, causes additional reperfusion injury. This study aimed to investigate the role of the ISR in myocardial ischemia/reperfusion (I/R).
METHODS
Cardiac-specific gain- and loss-of-function approaches for the ISR were used in vivo. Myocardial I/R was achieved by ligation of the cardiac left anterior descending artery for 45 minutes followed by reperfusion for different times. Cardiac function was assessed by echocardiography. Cultured H9c2 cells, primary rat cardiomyocytes, and mouse embryonic fibroblasts were used to dissect underlying molecular mechanisms. Tandem mass tag labeling and mass spectrometry was conducted to identify protein targets of the ISR. Pharmacologic means were tested to manipulate the ISR for therapeutic exploration.
RESULTS
We show that the PERK (PKR-like endoplasmic reticulum resident kinase)/eIF2α (α subunit of eukaryotic initiation factor 2) axis of the ISR is strongly induced by I/R in cardiomyocytes in vitro and in vivo. We further reveal a physiologic role of PERK/eIF2α signaling by showing that acute activation of PERK in the heart confers robust cardioprotection against reperfusion injury. In contrast, cardiac-specific deletion of PERK aggravates cardiac responses to reperfusion. Mechanistically, the ISR directly targets mitochondrial complexes through translational suppression. We identify NDUFAF2 (NADH:ubiquinone oxidoreductase complex assembly factor 2), an assembly factor of mitochondrial complex I, as a selective target of PERK. Overexpression of PERK suppresses the protein expression of NDUFAF2 and PERK inhibition causes an increase of NDUFAF2. Silencing of NDUFAF2 significantly rescues cardiac cell survival from PERK knockdown under I/R. We show that activation of PERK/eIF2α signaling reduces mitochondrial complex-derived reactive oxygen species and improves cardiac cell survival in response to I/R. Moreover, pharmacologic stimulation of the ISR protects the heart against reperfusion damage, even after the restoration of occluded coronary artery, highlighting clinical relevance for myocardial infarction treatment.
CONCLUSIONS
These results suggest that the ISR improves cell survival and mitigates reperfusion damage by selectively suppressing mitochondrial protein synthesis and reducing oxidative stress in the heart.
Topics: Animals; Humans; Mice; Mice, Knockout; Mitochondrial Proteins; Oxidative Stress; Protein Biosynthesis
PubMed: 34583519
DOI: 10.1161/CIRCULATIONAHA.120.053125 -
Signal Transduction and Targeted Therapy Apr 2023tsRNAs (tRNA-derived small RNAs), as products of the stress response, exert considerable influence on stress response and injury regulation. However, it remains largely...
tsRNAs (tRNA-derived small RNAs), as products of the stress response, exert considerable influence on stress response and injury regulation. However, it remains largely unclear whether tsRNAs can ameliorate liver injury. Here, we demonstrate the roles of tsRNAs in alleviating liver injury by utilizing the loss of NSun2 (NOP2/Sun domain family, member 2) as a tsRNAs-generating model. Mechanistically, the loss of NSun2 reduces methyluridine-U5 (mU) and cytosine-C5 (mC) of tRNAs, followed by the production of various tsRNAs, especially Class I tsRNAs (tRF-1s). Through further screening, we show that tRF-Gln-CTG-026 (tG026), the optimal tRF-1, ameliorates liver injury by repressing global protein synthesis through the weakened association between TSR1 (pre-rRNA-processing protein TSR1 homolog) and pre-40S ribosome. This study indicates the potential of tsRNA-reduced global protein synthesis in liver injury and repair, suggesting a potential therapeutic strategy for liver injury.
Topics: Protein Biosynthesis; Ribosomes; RNA; RNA Precursors; RNA Processing, Post-Transcriptional; Animals; Mice; Chemical and Drug Induced Liver Injury
PubMed: 37015921
DOI: 10.1038/s41392-023-01351-5 -
Journal of Molecular Cell Biology Oct 2019Most eukaryotic mRNAs are translated in a cap-dependent fashion; however, under stress conditions, the cap-independent translation driven by internal ribosomal entry... (Review)
Review
Most eukaryotic mRNAs are translated in a cap-dependent fashion; however, under stress conditions, the cap-independent translation driven by internal ribosomal entry sites (IRESs) can serve as an alternative mechanism for protein production. Many IRESs have been discovered from viral or cellular mRNAs to promote ribosome assembly and initiate translation by recruiting different trans-acting factors. Although the mechanisms of translation initiation driven by viral IRESs are relatively well understood, the existence of cellular IRESs is still under debate due to the limitations of translation reporter systems used to assay IRES activities. A recent screen identified > 1000 putative IRESs from viral and human mRNAs, expanding the scope and mechanism for cap-independent translation. Additionally, a large number of circular RNAs lacking free ends were identified in eukaryotic cells, many of which are found to be translated through IRESs. These findings suggest that IRESs may play a previously unappreciated role in driving translation of the new type of mRNA, implying a hidden proteome produced from cap-independent translation.
Topics: Humans; Internal Ribosome Entry Sites; Models, Biological; Protein Biosynthesis; Proteome; Ribosomes
PubMed: 31504667
DOI: 10.1093/jmcb/mjz091 -
Nature Sep 2021Single-cell sequencing methods have enabled in-depth analysis of the diversity of cell types and cell states in a wide range of organisms. These tools focus...
Single-cell sequencing methods have enabled in-depth analysis of the diversity of cell types and cell states in a wide range of organisms. These tools focus predominantly on sequencing the genomes, epigenomes and transcriptomes of single cells. However, despite recent progress in detecting proteins by mass spectrometry with single-cell resolution, it remains a major challenge to measure translation in individual cells. Here, building on existing protocols, we have substantially increased the sensitivity of these assays to enable ribosome profiling in single cells. Integrated with a machine learning approach, this technology achieves single-codon resolution. We validate this method by demonstrating that limitation for a particular amino acid causes ribosome pausing at a subset of the codons encoding the amino acid. Of note, this pausing is only observed in a sub-population of cells correlating to its cell cycle state. We further expand on this phenomenon in non-limiting conditions and detect pronounced GAA pausing during mitosis. Finally, we demonstrate the applicability of this technique to rare primary enteroendocrine cells. This technology provides a first step towards determining the contribution of the translational process to the remarkable diversity between seemingly identical cells.
Topics: Amino Acids; Animals; Cell Cycle; Cell Line; Codon; Female; Humans; Machine Learning; Male; Mice; Peptide Chain Elongation, Translational; Peptide Chain Initiation, Translational; Peptide Chain Termination, Translational; Protein Biosynthesis; RNA-Seq; Reproducibility of Results; Ribosomes; Single-Cell Analysis
PubMed: 34497418
DOI: 10.1038/s41586-021-03887-4