19 research outputs found
A SARS-CoV-2 protein interaction map reveals targets for drug repurposing
The novel coronavirus SARS-CoV-2, the causative agent of COVID-19 respiratory disease, has infected over 2.3 million people, killed over 160,000, and caused worldwide social and economic disruption1,2. There are currently no antiviral drugs with proven clinical efficacy, nor are there vaccines for its prevention, and these efforts are hampered by limited knowledge of the molecular details of SARS-CoV-2 infection. To address this, we cloned, tagged and expressed 26 of the 29 SARS-CoV-2 proteins in human cells and identified the human proteins physically associated with each using affinity-purification mass spectrometry (AP-MS), identifying 332 high-confidence SARS-CoV-2-human protein-protein interactions (PPIs). Among these, we identify 66 druggable human proteins or host factors targeted by 69 compounds (29 FDA-approved drugs, 12 drugs in clinical trials, and 28 preclinical compounds). Screening a subset of these in multiple viral assays identified two sets of pharmacological agents that displayed antiviral activity: inhibitors of mRNA translation and predicted regulators of the Sigma1 and Sigma2 receptors. Further studies of these host factor targeting agents, including their combination with drugs that directly target viral enzymes, could lead to a therapeutic regimen to treat COVID-19
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The SWI/SNF complex regulates splicing outcomes to determine cell fate in response to environmental cues in Saccharomyces cerevisiae
Despite its relatively streamlined genome, there are important examples of regulated RNA splicing in Saccharomyces cerevisiae. Here we show crucial roles for the chromatin remodeling complex SWI/SNF in splicing regulation in response to environmental changes. Nutrient-dependent downregulation of Snf2, the ATPase subunit of SWI/SNF, regulates downregulation of ribosomal protein genes (RPGs). RPGs are intron-enriched, and are highly transcribed. We show that their downregulation causes spliceosome redistribution from this abundant class of intron-containing RNAs to transcripts containing non-canonical splice-signals, which otherwise have poor affinity for the spliceosome. Meiosis in S. cerevisiae is a response to prolonged starvation, involving regulated transcription and splicing of meiosis-specific transcripts. Splicing of a subset of these relies upon the meiosis-specific splicing activator Mer1. We find that SWI/SNF affects meiotic splicing in multiple ways. First, meiosis-specific downregulation of Snf2 leads to RPG downregulation and spliceosome redistribution to Mer1-regulated transcripts. Secondly, Mer1 expression is SWI/SNF dependent—Snf2 is poised at the MER1 promoter, and timing of Snf2 downregulation in relation to acetylation states of both itself and its target genomic loci allows coordination between these mechanisms. Hence, the SWI/SNF complex directs regulated meiotic splicing in S. cerevisiae. Furthermore, Snf2 itself is subject to precise regulation in response to cellular needs via several novel modes of RNA processing and regulation, as well as control of protein acetylation and turnover. This multi-level coordinated regulation orchestrates activity and targets of splicing programs as a cellular adaptive strategy in response to environmental stresses. We also report roles for the SWI/SNF complex in respiration, partially via splicing regulation. Nutrient-dependent decrease in Snf2 leads to increase in PTC7 splicing, due to RPG downregulation and spliceosome redistribution. The spliced PTC7 transcript encodes a mitochondrial phosphatase regulator of Coenzyme Q6 (CoQ6) biosynthesis, a mitochondrial redox-active lipid essential for respiration, and increased PTC7 splicing increases CoQ6 levels. Contrastingly, the nonspliced PTC7 isoform encodes a protein repressing CoQ6 biosynthesis via as-yet-unknown mechanisms. These findings establish a novel role for SWI/SNF in the transition of yeast cells from fermentative to respiratory metabolism. Overall, the SWI/SNF complex regulates cellular stress responses by redirecting energy from translation to specialized splicing programs
The SWI/SNF complex regulates splicing outcomes to determine cell fate in response to environmental cues in Saccharomyces cerevisiae
Despite its relatively streamlined genome, there are important examples of regulated RNA splicing in Saccharomyces cerevisiae. Here we show crucial roles for the chromatin remodeling complex SWI/SNF in splicing regulation in response to environmental changes. Nutrient-dependent downregulation of Snf2, the ATPase subunit of SWI/SNF, regulates downregulation of ribosomal protein genes (RPGs). RPGs are intron-enriched, and are highly transcribed. We show that their downregulation causes spliceosome redistribution from this abundant class of intron-containing RNAs to transcripts containing non-canonical splice-signals, which otherwise have poor affinity for the spliceosome. Meiosis in S. cerevisiae is a response to prolonged starvation, involving regulated transcription and splicing of meiosis-specific transcripts. Splicing of a subset of these relies upon the meiosis-specific splicing activator Mer1. We find that SWI/SNF affects meiotic splicing in multiple ways. First, meiosis-specific downregulation of Snf2 leads to RPG downregulation and spliceosome redistribution to Mer1-regulated transcripts. Secondly, Mer1 expression is SWI/SNF dependent—Snf2 is poised at the MER1 promoter, and timing of Snf2 downregulation in relation to acetylation states of both itself and its target genomic loci allows coordination between these mechanisms. Hence, the SWI/SNF complex directs regulated meiotic splicing in S. cerevisiae. Furthermore, Snf2 itself is subject to precise regulation in response to cellular needs via several novel modes of RNA processing and regulation, as well as control of protein acetylation and turnover. This multi-level coordinated regulation orchestrates activity and targets of splicing programs as a cellular adaptive strategy in response to environmental stresses. We also report roles for the SWI/SNF complex in respiration, partially via splicing regulation. Nutrient-dependent decrease in Snf2 leads to increase in PTC7 splicing, due to RPG downregulation and spliceosome redistribution. The spliced PTC7 transcript encodes a mitochondrial phosphatase regulator of Coenzyme Q6 (CoQ6) biosynthesis, a mitochondrial redox-active lipid essential for respiration, and increased PTC7 splicing increases CoQ6 levels. Contrastingly, the nonspliced PTC7 isoform encodes a protein repressing CoQ6 biosynthesis via as-yet-unknown mechanisms. These findings establish a novel role for SWI/SNF in the transition of yeast cells from fermentative to respiratory metabolism. Overall, the SWI/SNF complex regulates cellular stress responses by redirecting energy from translation to specialized splicing programs
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The Traffic Jam: Polyamine Prevalence Pauses Protein Production.
In this issue of Molecular Cell, Ivanov et al. (2018) demonstrate how cells sense metabolite levels and regulate production of their biosynthetic enzymes through the formation of "ribosome queues.
DDX3X and DDX3Y are redundant in protein synthesis
DDX3 is a DEAD-box RNA helicase that regulates translation and is encoded by the X- and Y-linked paralogs DDX3X and DDX3Y While DDX3X is ubiquitously expressed in human tissues and essential for viability, DDX3Y is male-specific and shows lower and more variable expression than DDX3X in somatic tissues. Heterozygous genetic lesions in DDX3X mediate a class of developmental disorders called DDX3X syndrome, while loss of DDX3Y is implicated in male infertility. One possible explanation for female-bias in DDX3X syndrome is that DDX3Y encodes a polypeptide with different biochemical activity. In this study, we use ribosome profiling and in vitro translation to demonstrate that the X- and Y-linked paralogs of DDX3 play functionally redundant roles in translation. We find that transcripts that are sensitive to DDX3X depletion or mutation are rescued by complementation with DDX3Y. Our data indicate that DDX3X and DDX3Y proteins can functionally complement each other in the context of mRNA translation in human cells. DDX3Y is not expressed in a large fraction of the central nervous system. These findings suggest that expression differences, not differences in paralog-dependent protein synthesis, underlie the sex-bias of DDX3X-associated diseases
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The histone variant H2A.Z promotes efficient cotranscriptional splicing in S. cerevisiae
In eukaryotes, a dynamic ribonucleic protein machine known as the spliceosome catalyzes the removal of introns from premessenger RNA (pre-mRNA). Recent studies show the processes of RNA synthesis and RNA processing to be spatio-temporally coordinated, indicating that RNA splicing takes place in the context of chromatin. H2A.Z is a highly conserved histone variant of the canonical histone H2A. In Saccharomyces cerevisiae, H2A.Z is deposited into chromatin by the SWR-C complex, is found near the 5' ends of protein-coding genes, and has been implicated in transcription regulation. Here we show that splicing of intron-containing genes in cells lacking H2A.Z is impaired, particularly under suboptimal splicing conditions. Cells lacking H2A.Z are especially dependent on a functional U2 snRNP (small nuclear RNA [snRNA] plus associated proteins), as H2A.Z shows extensive genetic interactions with U2 snRNP-associated proteins, and RNA sequencing (RNA-seq) reveals that introns with nonconsensus branch points are particularly sensitive to H2A.Z loss. Consistently, H2A.Z promotes efficient spliceosomal rearrangements involving the U2 snRNP, as H2A.Z loss results in persistent U2 snRNP association and decreased recruitment of downstream snRNPs to nascent RNA. H2A.Z impairs transcription elongation, suggesting that spliceosome rearrangements are tied to H2A.Z's role in elongation. Depletion of disassembly factor Prp43 suppresses H2A.Z-mediated splice defects, indicating that, in the absence of H2A.Z, stalled spliceosomes are disassembled, and unspliced RNAs are released. Together, these data demonstrate that H2A.Z is required for efficient pre-mRNA splicing and indicate a role for H2A.Z in coordinating the kinetics of transcription elongation and splicing
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A ubiquitous GC content signature underlies multimodal mRNA regulation by DDX3X
The road from transcription to protein synthesis is paved with many obstacles, allowing for several modes of post-transcriptional regulation of gene expression. A fundamental player in mRNA biology is DDX3X, an RNA binding protein that canonically regulates mRNA translation. By monitoring dynamics of mRNA abundance and translation following DDX3X depletion, we observe stabilization of translationally suppressed mRNAs. We use interpretable statistical learning models to uncover GC content in the coding sequence as the major feature underlying RNA stabilization. This result corroborates GC content-related mRNA regulation detectable in other studies, including hundreds of ENCODE datasets and recent work focusing on mRNA dynamics in the cell cycle. We provide further evidence for mRNA stabilization by detailed analysis of RNA-seq profiles in hundreds of samples, including a Ddx3x conditional knockout mouse model exhibiting cell cycle and neurogenesis defects. Our study identifies a ubiquitous feature underlying mRNA regulation and highlights the importance of quantifying multiple steps of the gene expression cascade, where RNA abundance and protein production are often uncoupled
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Chromatin-remodeling SWI/SNF complex regulates coenzyme Q6 synthesis and a metabolic shift to respiration in yeast.
Despite its relatively streamlined genome, there are many important examples of regulated RNA splicing in Saccharomyces cerevisiae Here, we report a role for the chromatin remodeler SWI/SNF in respiration, partially via the regulation of splicing. We find that a nutrient-dependent decrease in Snf2 leads to an increase in splicing of the PTC7 transcript. The spliced PTC7 transcript encodes a mitochondrial phosphatase regulator of biosynthesis of coenzyme Q6 (ubiquinone or CoQ6) and a mitochondrial redox-active lipid essential for electron and proton transport in respiration. Increased splicing of PTC7 increases CoQ6 levels. The increase in PTC7 splicing occurs at least in part due to down-regulation of ribosomal protein gene expression, leading to the redistribution of spliceosomes from this abundant class of intron-containing RNAs to otherwise poorly spliced transcripts. In contrast, a protein encoded by the nonspliced isoform of PTC7 represses CoQ6 biosynthesis. Taken together, these findings uncover a link between Snf2 expression and the splicing of PTC7 and establish a previously unknown role for the SWI/SNF complex in the transition of yeast cells from fermentative to respiratory modes of metabolism