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Cell Nov 2023A fundamental feature of cellular growth is that total protein and RNA amounts increase with cell size to keep concentrations approximately constant. A key component of...
A fundamental feature of cellular growth is that total protein and RNA amounts increase with cell size to keep concentrations approximately constant. A key component of this is that global transcription rates increase in larger cells. Here, we identify RNA polymerase II (RNAPII) as the limiting factor scaling mRNA transcription with cell size in budding yeast, as transcription is highly sensitive to the dosage of RNAPII but not to other components of the transcriptional machinery. Our experiments support a dynamic equilibrium model where global RNAPII transcription at a given size is set by the mass action recruitment kinetics of unengaged nucleoplasmic RNAPII to the genome. However, this only drives a sub-linear increase in transcription with size, which is then partially compensated for by a decrease in mRNA decay rates as cells enlarge. Thus, limiting RNAPII and feedback on mRNA stability work in concert to scale mRNA amounts with cell size.
Topics: Cell Size; Feedback; RNA Polymerase II; RNA Stability; RNA, Messenger; Transcription, Genetic
PubMed: 37944513
DOI: 10.1016/j.cell.2023.10.012 -
Cell Jul 2023Readthrough into the 3' untranslated region (3' UTR) of the mRNA results in the production of aberrant proteins. Metazoans efficiently clear readthrough proteins, but...
Readthrough into the 3' untranslated region (3' UTR) of the mRNA results in the production of aberrant proteins. Metazoans efficiently clear readthrough proteins, but the underlying mechanisms remain unknown. Here, we show in Caenorhabditis elegans and mammalian cells that readthrough proteins are targeted by a coupled, two-level quality control pathway involving the BAG6 chaperone complex and the ribosome-collision-sensing protein GCN1. Readthrough proteins with hydrophobic C-terminal extensions (CTEs) are recognized by SGTA-BAG6 and ubiquitylated by RNF126 for proteasomal degradation. Additionally, cotranslational mRNA decay initiated by GCN1 and CCR4/NOT limits the accumulation of readthrough products. Unexpectedly, selective ribosome profiling uncovered a general role of GCN1 in regulating translation dynamics when ribosomes collide at nonoptimal codons, enriched in 3' UTRs, transmembrane proteins, and collagens. GCN1 dysfunction increasingly perturbs these protein classes during aging, resulting in mRNA and proteome imbalance. Our results define GCN1 as a key factor acting during translation in maintaining protein homeostasis.
Topics: Animals; Protein Biosynthesis; Ribosomes; Proteins; RNA, Messenger; Codon, Terminator; Mammals
PubMed: 37339632
DOI: 10.1016/j.cell.2023.05.035 -
RNA (New York, N.Y.) Apr 2024Epitranscriptomics refers to chemical changes in RNAs and includes numerous chemical types with varying stoichiometry and functions. RNA modifications are highly diverse...
Epitranscriptomics refers to chemical changes in RNAs and includes numerous chemical types with varying stoichiometry and functions. RNA modifications are highly diverse in chemistry and respond in cell-type- and cell-state-dependent manners that enable and facilitate the execution of a wide array of biological functions. This includes roles in the regulation of transcription, translation, chromatin maintenance, immune response, and many other processes. This special issue presents the past, present, and future of epitranscriptomics research with a focus on mRNA. It includes perspectives from experts in the field, with the goal of encouraging discussions and debates that will further advance this area of research and enable it to realize its full potential in basic research and applications to human health and disease.
Topics: Humans; RNA, Messenger; RNA; RNA Processing, Post-Transcriptional
PubMed: 38531649
DOI: 10.1261/rna.079993.124 -
Nature Cell Biology Nov 2023N-methyladenosine (mA) is the most abundant internal mRNA nucleotide modification in mammals, regulating critical aspects of cell physiology and differentiation. The...
N-methyladenosine (mA) is the most abundant internal mRNA nucleotide modification in mammals, regulating critical aspects of cell physiology and differentiation. The YTHDF proteins are the primary readers of mA modifications and exert physiological functions of mA in the cytosol. Elucidating the regulatory mechanisms of YTHDF proteins is critical to understanding mA biology. Here we report a mechanism that protein post-translational modifications control the biological functions of the YTHDF proteins. We find that YTHDF1 and YTHDF3, but not YTHDF2, carry high levels of nutrient-sensing O-GlcNAc modifications. O-GlcNAcylation attenuates the translation-promoting function of YTHDF1 and YTHDF3 by blocking their interactions with proteins associated with mRNA translation. We further demonstrate that O-GlcNAc modifications on YTHDF1 and YTHDF3 regulate the assembly, stability and disassembly of stress granules to enable better recovery from stress. Therefore, our results discover an important regulatory pathway of YTHDF functions, adding an additional layer of complexity to the post-transcriptional regulation function of mRNA mA.
Topics: Animals; Protein Processing, Post-Translational; Proteins; RNA, Messenger; Gene Expression Regulation; Mammals
PubMed: 37945829
DOI: 10.1038/s41556-023-01258-x -
Nature Nanotechnology Nov 2023Effective cancer immunotherapy is usually blocked by immunosuppressive factors in the tumour microenvironment, resulting in tumour promotion, metastasis and recurrence....
Effective cancer immunotherapy is usually blocked by immunosuppressive factors in the tumour microenvironment, resulting in tumour promotion, metastasis and recurrence. Here we combine lipid nanoparticle-mRNA formulations and dendritic cell therapy (named CATCH) to boost the cancer-immunity cycle via progressive steps to overcome the immunosuppressive tumour microenvironment. Multiple types of sugar-alcohol-derived lipid nanoparticles are conceived to modulate the cancer-immunity cycle. First, one type of lipid nanoparticle containing CD40 ligand mRNA induces robust immunogenic cell death in tumoural tissues, leading to the release of tumour-associated antigens and the expression of CD40 ligand. Next, dendritic cells engineered by another type of lipid nanoparticle encapsulating CD40 mRNA are adoptively transferred, which are then activated by the CD40 ligand molecules in tumoural tissues. This promotes the secretion of multiple cytokines and chemokines, and the upregulation of co-stimulatory molecules on dendritic cells, which are crucial for reprogramming the tumour microenvironment and priming the T-cell responses. After dendritic cells present tumour-associated antigens to T cells, all the above stepwise events contribute to boosting a potent tumour-specific T-cell immunity that eradicates established tumours, suppresses distal lesions and prevents tumour rechallenge.
Topics: Humans; CD40 Ligand; RNA, Messenger; Dendritic Cells; Neoplasms; Tumor Microenvironment
PubMed: 37500773
DOI: 10.1038/s41565-023-01453-9 -
Cell Sep 2023Cellular homeostasis requires the robust control of biomolecule concentrations, but how do millions of mRNAs coordinate their stoichiometries in the face of dynamic...
Cellular homeostasis requires the robust control of biomolecule concentrations, but how do millions of mRNAs coordinate their stoichiometries in the face of dynamic translational changes? Here, we identified a two-tiered mechanism controlling mRNA:mRNA and mRNA:protein stoichiometries where mRNAs super-assemble into condensates with buffering capacity and sorting selectivity through phase-transition mechanisms. Using C. elegans oogenesis arrest as a model, we investigated the transcriptome cytosolic reorganization through the sequencing of RNA super-assemblies coupled with single mRNA imaging. Tightly repressed mRNAs self-assembled into same-sequence nanoclusters that further co-assembled into multiphase condensates. mRNA self-sorting was concentration dependent, providing a self-buffering mechanism that is selective to sequence identity and controls mRNA:mRNA stoichiometries. The cooperative sharing of limiting translation repressors between clustered mRNAs prevented the disruption of mRNA:repressor stoichiometries in the cytosol. Robust control of mRNA:mRNA and mRNA:protein stoichiometries emerges from mRNA self-demixing and cooperative super-assembly into multiphase multiscale condensates with dynamic storage capacity.
Topics: Animals; Caenorhabditis elegans; Oogenesis; Protein Biosynthesis; RNA Transport; RNA, Messenger; Proteins; Biomolecular Condensates
PubMed: 37703874
DOI: 10.1016/j.cell.2023.08.018 -
Nature Communications Oct 2023N-acetyltransferase 10 (NAT10) is an N-acetylcytidine (acC) writer that catalyzes RNA acetylation at cytidine N position on tRNAs, rRNAs and mRNAs. Recently, NAT10 and...
N-acetyltransferase 10 (NAT10) is an N-acetylcytidine (acC) writer that catalyzes RNA acetylation at cytidine N position on tRNAs, rRNAs and mRNAs. Recently, NAT10 and the associated acC have been reported to increase the stability of HIV-1 transcripts. Here, we show that NAT10 catalyzes acC addition to the polyadenylated nuclear RNA (PAN), a long non-coding RNA encoded by the oncogenic DNA virus Kaposi's sarcoma-associated herpesvirus (KSHV), triggering viral lytic reactivation from latency. Mutagenesis of acC sites in PAN RNA in the context of KSHV infection abolishes PAN acC modifications, downregulates the expression of viral lytic genes and reduces virion production. NAT10 knockdown or mutagenesis erases acC modifications of PAN RNA and increases its instability, and prevents KSHV reactivation. Furthermore, PAN acC modification promotes NAT10 recruitment of IFN-γ-inducible protein-16 (IFI16) mRNA, resulting in its acC acetylation, mRNA stability and translation, and eventual inflammasome activation. These results reveal a novel mechanism of viral and host acC modifications and the associated complexes as a critical switch of KSHV replication and antiviral immunity.
Topics: Herpesvirus 8, Human; Inflammasomes; RNA, Messenger; RNA, Nuclear; Cytidine; RNA Stability; Virus Replication; Gene Expression Regulation, Viral
PubMed: 37816771
DOI: 10.1038/s41467-023-42135-3 -
Cell Death and Differentiation Jul 2023The mitochondrial transmembrane (TMEM) protein family has several essential physiological functions. However, its roles in cardiomyocyte proliferation and cardiac...
The mitochondrial transmembrane (TMEM) protein family has several essential physiological functions. However, its roles in cardiomyocyte proliferation and cardiac regeneration remain unclear. Here, we detected that TMEM11 inhibits cardiomyocyte proliferation and cardiac regeneration in vitro. TMEM11 deletion enhanced cardiomyocyte proliferation and restored heart function after myocardial injury. In contrast, TMEM11-overexpression inhibited neonatal cardiomyocyte proliferation and regeneration in mouse hearts. TMEM11 directly interacted with METTL1 and enhanced mG methylation of Atf5 mRNA, thereby increasing ATF5 expression. A TMEM11-dependent increase in ATF5 promoted the transcription of Inca1, an inhibitor of cyclin-dependent kinase interacting with cyclin A1, which suppressed cardiomyocyte proliferation. Hence, our findings revealed that TMEM11-mediated mG methylation is involved in the regulation of cardiomyocyte proliferation, and targeting the TMEM11-METTL1-ATF5-INCA1 axis may serve as a novel therapeutic strategy for promoting cardiac repair and regeneration.
Topics: Animals; Mice; Cell Proliferation; Methylation; Myocytes, Cardiac; Protein Processing, Post-Translational; RNA, Messenger
PubMed: 37286744
DOI: 10.1038/s41418-023-01179-0 -
The AAPS Journal Oct 2023Delivery of RNA using nanomaterials has emerged as a new modality to expand therapeutic applications in biomedical research. However, the delivery of RNA presents unique... (Review)
Review
Delivery of RNA using nanomaterials has emerged as a new modality to expand therapeutic applications in biomedical research. However, the delivery of RNA presents unique challenges due to its susceptibility to degradation and the requirement for efficient intracellular delivery. The integration of nanotechnologies with RNA delivery has addressed many of these challenges. In this review, we discuss different strategies employed in the design and development of nanomaterials for RNA delivery. We also highlight recent advances in the pharmaceutical applications of RNA delivered via nanomaterials. Various nanomaterials, such as lipids, polymers, peptides, nucleic acids, and inorganic nanomaterials, have been utilized for delivering functional RNAs, including messenger RNA (mRNA), small interfering RNA, single guide RNA, and microRNA. Furthermore, the utilization of nanomaterials has expanded the applications of functional RNA as active pharmaceutical ingredients. For instance, the delivery of antigen-encoding mRNA using nanomaterials enables the transient expression of vaccine antigens, leading to immunogenicity and prevention against infectious diseases. Additionally, nanomaterial-mediated RNA delivery has been investigated for engineering cells to express exogenous functional proteins. Nanomaterials have also been employed for co-delivering single guide RNA and mRNA to facilitate gene editing of genetic diseases. Apart from the progress made in RNA medicine, we discuss the current challenges and future directions in this field.
Topics: Nanomedicine; Nanotechnology; Pharmaceutical Preparations; RNA, Small Interfering; RNA, Messenger
PubMed: 37784005
DOI: 10.1208/s12248-023-00860-z -
Molecular Cell Jul 2023Co-transcriptional capping of the nascent pre-mRNA 5' end prevents degradation of RNA polymerase (Pol) II transcripts and suppresses the innate immune response. Here, we...
Co-transcriptional capping of the nascent pre-mRNA 5' end prevents degradation of RNA polymerase (Pol) II transcripts and suppresses the innate immune response. Here, we provide mechanistic insights into the three major steps of human co-transcriptional pre-mRNA capping based on six different cryoelectron microscopy (cryo-EM) structures. The human mRNA capping enzyme, RNGTT, first docks to the Pol II stalk to position its triphosphatase domain near the RNA exit site. The capping enzyme then moves onto the Pol II surface, and its guanylyltransferase receives the pre-mRNA 5'-diphosphate end. Addition of a GMP moiety can occur when the RNA is ∼22 nt long, sufficient to reach the active site of the guanylyltransferase. For subsequent cap(1) methylation, the methyltransferase CMTR1 binds the Pol II stalk and can receive RNA after it is grown to ∼29 nt in length. The observed rearrangements of capping factors on the Pol II surface may be triggered by the completion of catalytic reaction steps and are accommodated by domain movements in the elongation factor DRB sensitivity-inducing factor (DSIF).
Topics: Humans; RNA Processing, Post-Transcriptional; RNA, Messenger; Cryoelectron Microscopy; RNA Polymerase II; Transcription, Genetic; Methyltransferases; Models, Chemical
PubMed: 37369200
DOI: 10.1016/j.molcel.2023.06.002