VAP-A and its binding partner CERT drive biogenesis of RNA-containing extracellular vesicles at ER membrane contact sites
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA777941 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR16775346.fastq.gz | Fastqsanger.gz | |||
| SRR16775347.fastq.gz | Fastqsanger.gz | |||
| SRR16775348.fastq.gz | Fastqsanger.gz | |||
| SRR16775349.fastq.gz | Fastqsanger.gz | |||
| SRR16775350.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 27 |
Similar Datasets
Project description:RNA transfer via extracellular vesicles (EVs) influences cell phenotypes; however, lack of information regarding biogenesis of RNA-containing EVs has limited progress in the field. Here, we identify endoplasmic reticulum membrane contact sites (ER MCS) as platforms for generation of RNA-containing EVs. We identify a subpopulation of small EVs that is highly enriched in RNA and regulated by the ER MCS linker protein VAP-A. Functionally, VAP-A-regulated EVs are critical for miR-100 transfer between cells and in vivo tumor formation. Lipid analysis of VAP-A-knockdown EVs revealed reductions in the EV biogenesis lipid ceramide. Knockdown of the VAP-A-binding ceramide transfer protein CERT led to similar defects in EV RNA content. Imaging experiments revealed that VAP-A promotes lumenal filling of multivesicular bodies (MVBs). CERT localizes to MVBs, and the ceramide-generating enzyme neutral sphingomyelinase 2 colocalizes with VAP-A-positive ER. We propose that ceramide transfer via VAP-A-CERT linkages drives biogenesis of a select RNA-containing population.
2022-04-05 | GSE188223 | GEO
Project description:RNA transfer via extracellular vesicles (EVs) influences cell phenotypes; however, lack of information regarding biogenesis of RNA-containing EVs has limited progress in the field. Here, we identify endoplasmic reticulum membrane contact sites (ER MCSs) as platforms for the generation of RNA-containing EVs. We identify a subpopulation of small EVs that is highly enriched in RNA and regulated by the ER MCS linker protein VAP-A. Functionally, VAP-A-regulated EVs are critical for miR-100 transfer between cells and in vivo tumor formation. Lipid analysis of VAP-A-knockdown EVs revealed reductions in the EV biogenesis lipid ceramide. Knockdown of the VAP-A-binding ceramide transfer protein CERT led to similar defects in EV RNA content. Imaging experiments revealed that VAP-A promotes luminal filling of multivesicular bodies (MVBs), CERT localizes to MVBs, and the ceramide-generating enzyme neutral sphingomyelinase 2 colocalizes with VAP-A-positive ER. We propose that ceramide transfer via VAP-A-CERT linkages drives the biogenesis of a select RNA-containing EV population.
| S-EPMC9075344 | biostudies-literature
Project description:Membrane contact sites are cellular structures that mediate inter-organelle exchange and communication. The endoplasmic reticulum (ER), which forms a network extending throughout the cytoplasm, possesses two known major tether proteins, named VAP-A and VAP-B, that allow its interaction with other organelles. VAP-A and VAP-B interact with proteins from other organelles that possess a small VAP-interacting motif, named FFAT (two phenylalanines in an acidic track), and consequently scaffold contact sites implicating the ER. In this study, by using an unbiased proteomic approach, we identified a novel ER tether named MOtile SPerm Domain-containing protein 2 (MOSPD2). We showed that MOSPD2 possesses a Motile Sperm Protein (MSP) domain which binds FFAT motifs and consequently allows membrane tethering in vitro. Accordingly, MOSPD2, which is an ER-anchored protein, interacts with several FFAT-containing proteins bound to diverse organelles,such as endosomes, mitochondria or Golgi. Consequently, the specific interaction between MOSPD2 and these organelle-bound proteins results in the formation of contact sites between the ER and these organelles. Thus, MOSPD2 is a novel tether for membrane contact sites between the ER and the other cellular organelles.
2018-06-05 | PXD009575 | Pride
Project description:In response to cholesterol deprivation, SCAP escorts SREBP transcription factors from the endoplasmic reticulum to the Golgi complex for their proteolytic activation, leading to gene expression for cholesterol synthesis and uptake. Here, we show that in cholesterol-fed cells, ER-localized SCAP interacts through Sac1 phosphatidylinositol 4-phosphate (PI4P) phosphatase with a VAP-OSBP complex, which mediates counter-transport of ER cholesterol and Golgi PI4P at ER-Golgi membrane contact sites (MCSs). SCAP knockdown inhibited the turnover of PI4P, perhaps due to a cholesterol transport defect, and altered the subcellular distribution of the VAP-OSBP complex. As in the case of perturbation of lipid transfer complexes at ER-Golgi MCSs, SCAP knockdown inhibited the biogenesis of the trans-Golgi network-derived transport carriers CARTS, which was reversed by expression of wild-type SCAP or a Golgi transport-defective mutant, but not of cholesterol sensing-defective mutants. Altogether, our findings reveal a new role for SCAP under cholesterol-fed conditions in the facilitation of CARTS biogenesis via ER-Golgi MCSs, depending on the ER cholesterol.
| S-EPMC7654440 | biostudies-literature
Project description:VAP-A regulates extracellular vesicle biogenesis at ER-membrane contact sites. Proteomic analysis of purified small extracellular vesicles from control (Sc) and VAP-A-KD (KD1 and KD2) DKs-8 cells were conducted.
2023-11-10 | PXD043796 | Pride
Project description:The causative agent of Legionnaires’ disease, Legionella pneumophila, governs interactions with host cells by secreting ca. 330 different “effector” proteins. The facultative intracellular bacteria replicate in macrophages and amoeba within a unique compartment, the Legionella-containing vacuole (LCV). Hallmarks of LCV formation are the phosphoinositide (PI) lipid conversion from PtdIns(3)P to PtdIns(4)P, fusion with endoplasmic reticulum (ER)-derived vesicles and a tight association with the ER. Proteomics of purified LCVs revealed the presence of membrane contact sites (MCS) proteins implicated in lipid exchange. Using dually fluorescence-labeled Dictyostelium discoideum amoeba, we reveal that the VAMP-associated protein (Vap), the PtdIns(4)P 4-phosphatase Sac1, and the large fusion GTPase Sey1/atlastin-3 localize to the ER, but not to the LCV membrane, and these ER-resident proteins promote intracellular replication of L. pneumophila and LCV remodeling. Moreover, oxysterol binding proteins (OSBPs) exclusively localize to the ER (OsbH) or the LCV membrane (OsbK), respectively, and promote (OsbH) or restrict (OsbK) intracellular replication of L. pneumophila and LCV expansion. Furthermore, the PtdIns(4)P-subverting L. pneumophila effectors LepB and SidC also promote LCV remodeling. Taken together, the Legionella-driven PtdIns(4)P gradient at LCV-ER MCSs promotes Vap-, OSBP- and Sac1-dependent pathogen vacuole remodeling.
2023-02-21 | PXD034490 | Pride
Project description:The endoplasmic reticulum (ER) supports biosynthesis of proteins with diverse transmembrane domain (TMD) lengths and hydrophobicity. Features in transmembrane domains such as charged residues in ion channels are often functionally important, but could pose a challenge during cotranslational membrane insertion and folding. Our systematic proteomic approaches in both yeast and human cells revealed that the ER membrane protein complex (EMC) binds to and promotes the biogenesis of a range of multipass transmembrane proteins, with a particular enrichment for transporters. Proximity-specific ribosome profiling demonstrates that the EMC engages clients cotranslationally and immediately following clusters of TMDs enriched for charged residues. The EMC can remain associated after completion of translation, which both protects clients from premature proteasomal degradation and allows recruitment of substrate-specific and general chaperones. Thus, the EMC broadly enables the biogenesis of multipass transmembrane proteins containing destabilizing features, thereby mitigating the trade-off between function and stability.
2018-05-29 | GSE112891 | GEO
Project description:Membrane contact sites (MCS) are intracellular regions where two organelles come closer to exchange information and material. The majority of the endoplasmic reticulum (ER) MCS are attributed to the ER-localized tether proteins VAPA, VAPB, and MOSPD2. These recruit other proteins to the ER by interacting with their FFAT motifs. Here, we describe MOSPD1 and MOSPD3 as ER-localized tethers interacting with FFAT motif-containing proteins. Using BioID, we identify proteins interacting with VAP and MOSPD proteins and find that MOSPD1 and MOSPD3 prefer unconventional FFAT-related FFNT (two phenylalanines [FF] in a neutral tract) motifs. Moreover, VAPA/VAPB/MOSPD2 and MOSPD1/MOSPD3 assemble into two separate ER-resident complexes to interact with FFAT and FFNT motifs, respectively. Because of their ability to interact with FFNT motifs, MOSPD1 and MOSPD3 could form MCS between the ER and other organelles. Collectively, these findings expand the VAP family of proteins and highlight two separate complexes in control of interactions between intracellular compartments.
2023-06-29 | PXD043399 | Pride
Project description:The double-membrane-bound autophagosome is formed by the closure of a structure called the phagophore, origin of which is still unclear. The endoplasmic reticulum (ER) is clearly implicated in autophagosome biogenesis due to the presence of the omegasome subdomain positive for DFCP1, a phosphatidyl-inositol-3-phosphate (PI3P) binding protein. Contribution of other membrane sources, like the plasma membrane (PM), is still difficult to integrate in a global picture. Here we show that ER-plasma membrane contact sites are mobilized for autophagosome biogenesis, by direct implication of the tethering extended synaptotagmins (E-Syts) proteins. Imaging data revealed that early autophagic markers are recruited to E-Syt-containing domains during autophagy and that inhibition of E-Syts expression leads to a reduction in autophagosome biogenesis. Furthermore, we demonstrate that E-Syts are essential for autophagy-associated PI3P synthesis at the cortical ER membrane via the recruitment of VMP1, the stabilizing ER partner of the PI3KC3 complex. These results highlight the contribution of ER-plasma membrane tethers to autophagosome biogenesis regulation and support the importance of membrane contact sites in autophagy.
| S-EPMC5509996 | biostudies-literature
Project description:PIWI proteins and their associated small RNAs called PIWI-interacting RNAs (piRNAs) restrict transposon activity in animal gonads to ensure fertility. Distinct biogenesis pathways load piRNAs into the PIWI proteins MILI and MIWI2 in the mouse male embryonic germline. While most of MILI piRNAs derive via a slicer-independent pathway, a MILI slicer endonuclease-initiated pathway loads nuclear MIWI2 with a series of phased piRNAs. Tudor domain-containing 12 (TDRD12) and its interaction partner Exonuclease domain-containing 1 (EXD1) are required for loading MIWI2, but only Tdrd12 is essential for fertility, leaving us with no explanation for the physiological role of Exd1. Using an artificial piRNA precursor, we demonstrate that MILI-triggered piRNA biogenesis is greatly reduced in the Exd1 mutant. The situation deteriorates in the sensitized Exd1 mutant (Exd1-/-; Tdrd12+/-), where diminished MIWI2 piRNA levels de-repress LINE1 retrotransposons, causing infertility. Thus, EXD1 enhances slicing-triggered MIWI2 piRNA biogenesis via a functional interaction with TDRD12.
2018-09-27 | GSE119447 | GEO