FUS facilitates gene body export from nuclear condensates
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA975594 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR24716300.fastq.gz | Fastqsanger.gz | |||
| SRR24716301.fastq.gz | Fastqsanger.gz | |||
| SRR24716284.fastq.gz | Fastqsanger.gz | |||
| SRR24716285.fastq.gz | Fastqsanger.gz | |||
| SRR24716286.fastq.gz | Fastqsanger.gz |
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Project description:Membraneless nuclear condensates are an emerging model of transcriptional regulation that involves spatially clustered enhancers and genes forming transcription condensates (TCs). We used a physical isolation approach and DNA/nascent RNA analysis to investigate the global organization of the genome in TCs. Comparative analysis of the two revealed the dynamic nature of the genome, with stable localization of promoters, dynamic recruitment of enhancers, and dynamic export of gene bodies out of TCs upon gene activation. Our findings also suggest that the RNA binding protein FUS undergoes decondensation upon nascent RNA binding, facilitating gene body export out of TCs.
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Project description:Biomolecular condensates formed by phase separation offer a confined space where protein-protein interactions (PPIs) facilitate distinct biochemical reactions. As their aberrant behavior is associated with disease, it is crucial to understand the molecular organization of PPIs within condensates. Using quantitative time-resolved crosslinking mass spectrometry (XL-MS) we directly and specifically monitor PPIs and protein dynamics inside condensates formed by the protein fused in sarcoma (FUS) and identify its folded RNA recognition motif (RRM) as a key player of aberrant molecular aging. We find that the chaperone HspB8, but not a disease-associated mutant, prevents FUS aging. In XL-MS and partitioning experiments we find that the disordered region of HspB8 directs its α-crystallin domain (αCD) into FUS droplets where HspB8 prevents aging via condensate-specific αCD–RRM interactions. We propose that chaperones like HspB8 prevent aberrant phase transitions by stabilizing aggregation prone folded domains inside condensates in times of cellular stress.
2021-10-13 | PXD021115 | Pride
Project description:The Hippo pathway plays important roles in organ development, tissue homeostasis, and tumor growth, and its downstream effector TAZ is a transcriptional coactivator that promotes target gene expression through the formation of biomolecular condensates. However, the mechanisms that regulate the biophysical properties of TAZ condensates to enable Hippo signaling are not well understood. Here, using chemical cross-linking combined with an unbiased proteomic approach, we show that FUS associates with TAZ condensates and exerts a chaperone-like effect to maintain their proper liquidity and robust- transcriptional activity. Mechanistically, the low complexity sequence domain of FUS targets the coiled-coil domain of TAZ in a phosphorylation-regulated manner, which ensures the liquidity and dynamicity of TAZ condensates. In cells lacking FUS, TAZ condensates transition into gel/solid-like assembles with immobilized TAZ, leading to reduced expression of target genes and inhibition of pro-tumorigenic activity. Thus, our findings identify a chaperone-like function of FUS in Hippo regulation and demonstrate that appropriate biophysical properties of transcriptional condensates are essential for gene activation.
2023-02-17 | PXD040257 |
Project description:Biomolecular condensates formed by phase separation offer a confined space where protein-protein interactions (PPIs) facilitate distinct biochemical reactions. As their aberrant behavior is increasingly associated with disease, it is crucial to understand the molecular organization of PPIs within condensates. Using quantitative and time-resolved crosslinkingmass spectrometry we directly and specifically monitor PPIs and protein dynamics of fused in sarcoma (FUS) condensates and identify its RNA recognition motif (RRM) as a key player of condensate-specific PPIs and aging. We find that the chaperone HspB8, but not a disease-associated mutant, prevents FUS aging. The disordered region of HspB8 directs its α-crystallin domain (αCD) into FUS condensates. Here, RRM unfolding drives aggregation but binding of HspB8 via a droplet-specific αCD – RRM interface significantly delays the onset of aging. We propose that chaperones such as HspB8 prevent aberrant phase transitions by stabilizing aggregation prone RNA binding domains in times of cellular stress.
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Project description:FMRP facilitates the nuclear export of N6-methyladenosine-containing mRNAs.
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Project description:Stress Granules (SG) formation is a cellular protection mechanism, constituting a storage for untranslated mRNAs and RNA-binding proteins (RBPs); however, these condensates can turn into pathological aggregates, related to the onset of neurodegenerative diseases like Amyotrophic Lateral Sclerosis (ALS). This transition towards cytotoxic inclusions is triggered by ALS-causative mutations in the RBP FUS, which lead to its cytoplasmic mis-localization and accumulation in SG. Here, we describe the SG transcriptome in a neural context and describe several features for RNA recruitment in SG. We demonstrate that SG dynamics and RNA content are strongly modified by the incorporation of mutant FUS, switching to a more unstructured, AU-rich SG transcriptome. Moreover, we show that mutant FUS, together with its protein interactors and their target RNAs, are responsible for the reshaping of the mutant SG transcriptome with alterations that can be linked to neurodegeneration. Therefore, our data give a comprehensive view of the molecular differences between physiological and pathological SG in ALS conditions, showing how FUS mutations impact the RNA and protein population of these condensates.
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Project description:Biosynthesis of the bacterial flagellum depends on a flagellar-specific type III secretion system (T3SS), a protein export machine homologous to the export machinery of the virulence-associated injectisome. Six cytoplasmic (FliH/I/J/G/M/N) and seven integral membrane proteins (FlhA/B FliF/O/P/Q/R) form the flagellar basal body and are involved in transport of flagellar building blocks across the inner membrane in a protein motive force-dependent manner. FlhA/B and FliP/Q/R are conserved in all T3SS and form the export gate complex. FliO is absent in some T3SS and was presumed to regulate FliP during flagellum assembly. However, the molecular function of FliO and how the large, multi-component transmembrane export gate complex assembles in a coordinated manner remained enigmatic. Here, we demonstrate that FliO protects FliP from proteolytic degradation and promotes formation of FliP-containing subcomplexes during the first step of core export apparatus assembly. We further reveal the sub-cellular localization of FliO by dSTORM superresolution microscopy and show that FliO is not part of the assembled flagellar basal body. In summary, our results suggest that FliO functions as a novel, flagellar T3SS-specific chaperone, which facilitates quality control and productive assembly of the core T3SS export machinery.
2017-08-22 | PXD005597 | Pride
Project description:A heterochromatin-specific RNA export pathway facilitates piRNA production
| PRJNA522430 | ENA
Project description:Samples 1012398_F1-F8: Using recombinant purified mCherry-tagged FUS-IDR we reconstituted condensates in a soluble nuclear extract isolated from U2OS cells. The extract was fractionated by centrifugation into a supernatant and pellet. Labels 126, 127, 128 are supernatant replicates and labels 129, 130, 131 are pellet replicates.
Samples 994350_F1-8: Using recombinant purified mEGFP-tagged MED1-IDR we reconstituted condensates in a soluble nuclear extract isolated from U2OS cells. The extract was fractionated by centrifugation into a supernatant and pellet. Labels 126, 127, 128 are supernatant replicates and labels 129, 130, 131 are pellet replicates.
For the proximity biotinylation method, we used SILAC (Lys-6, Arg-6) in the media of cells expressing the control while both FUS-IDR and MED1-IDR cells were grown in natural isotope conditions. Cells were subject to the biotinylation protocol followed by nuclear extract preparation. Light and heavy extracts were mixed at a 1:1 ratio followed by streptavidin pulldown, stringent washing, and elution. Two independent replicates of SILAC data were collected for both FUS-IDR (QEX2_1011001 and ECL1_1073837) and MED1-IDR (QEX2_996337 and ECL1_1078014).
2023-11-28 | MSV000093520 | MassIVE
Project description:Myogenesis is a highly orchestrated process whereby muscle precursor cells, myoblasts, develop into muscle fibers to form skeletal muscle during embryogenesis and regenerate adult muscle. Here, we studied the RNA-binding protein FUS (fused in sarcoma), which has been implicated in muscular and neuromuscular pathologies but is poorly characterized in myogenesis. Given that FUS levels declined in human and mouse models of skeletal myogenesis, and that silencing FUS enhanced myogenesis, we hypothesized that FUS might be a repressor of myogenic differentiation. Interestingly, overexpression of FUS delayed myogenesis, accompanied by slower production of muscle differentiation markers. To identify the mechanisms through which FUS inhibits myogenesis, we uncovered RNA targets of FUS by ribonucleoprotein immunoprecipitation (RIP) followed by RNA-sequencing (RNA-seq) analysis. Stringent selection of the bound transcripts uncovered Tnnt1 mRNA, encoding troponin T (TNNT1), as a major effector of FUS influence on myogenesis. We found that in myoblasts, FUS sequestered Tnnt1 mRNA in the nucleus, preventing TNNT1 expression; however, reduction of FUS during myogenesis or by silencing FUS released Tnnt1 mRNA for export to the cytoplasm, where it accumulated and was translated into TNNT1, promoting myogenesis. We propose that FUS inhibits myogenesis by suppressing TNNT1 expression through a mechanism of nuclear mRNA retention.
2024-09-13 | GSE267276 | GEO