Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Integrative structural Interactomics reveals protein organization and structure in a giant virus

ABSTRACT: Giant viruses are large DNA viruses that infect unicellular and multicellular eukaryotes and form exceptionally large extracellular particles. (Meta)genomics and (meta)transcriptomics have provided insight into their diverse coding repertoire, but many of the proteins remain to be characterized as they lack homology with known proteins. Here, we integrated cross-linking mass spectrometry, quantitative proteomics, computational tools and cryo-EM data to characterize the protein architecture of intact Melbournevirus particles. Based on this, we allocated 88 viral proteins to different virion sub-compartments and proposed topology of 25 inner membrane proteins. We assigned eight components of the capsid in cryo-EM data, including proteins that tether the capsid shell to the membrane, reflecting key points in virion maturation. The data provide a valuable resource and demonstrate the power of an integrative approach to gain system-level structural insights into a poorly characterized biological system.

INSTRUMENT(S):

ORGANISM(S): Melbournevirus Acanthamoeba Castellanii Str. Neff

SUBMITTER: Lars Mühlberg

LAB HEAD: Fan Liu

PROVIDER: PXD064933 | Pride | 2026-05-20

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	CSMs_1FDR_R1.csv	Csv
	CSMs_1FDR_R2.csv	Csv
	CSMs_1FDR_R3.csv	Csv
	F1_20230509_LM_46_MELV_vir1_MeOH_20230510153630.raw	Raw
	F1_20230509_LM_46_MELV_vir2_MeOH.raw	Raw

Items per page:

1 - 5 of 160

Similar Datasets

Spatial, Quantitative and Functional Deconstruction of Virus and Host Protein Interactions Inside Intact Cytomegalovirus Particles

Project description:Viral infections involve packaging of the viral genome and proteins into virions. Knowing virion composition and structure is critical to understand viral pathogenesis. However, an integrated picture of virion proteome organization of large viruses, such as Herpesviruses, is lacking. Here we use cross-linking mass spectrometry to derive a spatially resolved structural interactome of intact human cytomegalovirus virions. We capture interactions of 82 host and 33 viral proteins, allowing us to de novo allocate them into distinct virion-layers and identify several host proteins as constitutive virion components recruited via specific protein interactions. The abundant viral protein pp150 forms domain-specific interactions with all virion layers and scaffolds numerous viral and host proteins such as PP1 phosphatase and 14-3-3 proteins, which are incorporated via nearby short linear motifs in pp150’s C-terminus. PP1 recruitment antagonizes 14-3-3 proteins and is pivotal during early and late viral replication steps. Collectively, this study gives a spatial and quantitative inventory of the system-wide organization and functional relevance of protein interactions inside native herpesvirus particles.

2023-06-14 | PXD031911 | Pride

Structural Host Virus Interactome Profiling of Intact Infected Cells

Project description:Virus-host protein-protein interactions (PPIs) are fundamental to viral infections, yet high-resolution identification within the native context of intact infected cells has remained an unsolved challenge. Here, we developed structural host-virus interactome profiling (SHVIP) that combines in situ crosslinking mass spectrometry with the enrichment of newly synthesized viral proteins from infected cells. SHVIP captures PPIs across intracellular compartments and at the intact host endomembrane system. It resolves PPIs to the protein domain level and seamlessly integrates with AlphaFold-based structural modeling, facilitating detailed predictions of PPI sites within structured and intrinsically disordered regions. We show that SHVIP captures parts of the virus-host PPI space that are elusive to traditional interaction proteomics approaches. By selectively disrupting several newly identified virus-host PPIs, we confirm SHVIP’s ability to uncover genuine virus-host PPIs in the intact complex environment of infected cells.

2025-06-13 | PXD047422 | Pride

Interaction of Human Cytomegalovirus pUL52 with major components of the viral DNA encapsidation network underlines its essential role in genome cleavage-packaging

Project description:Cleavage of human cytomegalovirus (HCMV) genomes and their packaging into capsids requires at least seven essential viral proteins, yet it is not completely understood how these proteins cooperate to accomplish this task. Besides the portal protein pUL104 and the terminase subunits pUL51, pUL56, and pUL89, the UL52 protein is also necessary for HCMV genome encapsidation, however, knowledge about pUL52 is scant. In the absence of pUL52, viral concatemers are not cleaved into unit-length genomes and no DNA-filled capsids are observed, yet no viral or cellular proteins interacting with pUL52 have been identified that would explain how pUL52 exerts its essential role in the HCMV infection cycle. In this study, we aimed at a comprehensive definition of pUL52-interacting proteins in infected cells. Using suitable HCMV mutants, we employed three complementary state-of-the-art proteomic approaches, namely biotin ligase-dependent proximity labeling, affinity purification and cross-linking mass spectrometry. These experiments, combined with thorough validation by immunoblotting, pointed to several viral DNA-associated proteins and key players pivotal for genome encapsidation as interactors of pUL52. The most noticeable direct pUL52 interaction partners were the terminase subunits pUL56 and pUL89 as well as the portal protein pUL104. Hence, we suggest a model of pUL52 function in which pUL52 mediates association of HCMV genomes with the terminase subunits and the capsid portal. Taken together, our data contribute to the understanding of an essential viral process previously recognized as a prominent antiviral target. Disturbing the identified pUL52 interactions may provide a starting point to develop novel antiviral medication.

2025-02-17 | PXD060825 | Pride

Yeast TLDc domain-containing proteins control assembly and subcellular localization of the V-ATPase

Project description:Yeast vacuoles, equivalent to lysosomes in other eukaryotes, are important acidic degradative organelles as well as storage compartments and signaling hubs. To perform these functions, they rely on important protein complexes, including the V-ATPase, responsible for organelle acidification. In this study, we used cross-linking mass spectrometry to characterize the protein complexes of isolated vacuoles. We were able to detect many known protein-protein interactions, including known protein complexes, as well as undescribed ones. Among these, we identified the uncharacterized TLDc domain-containing protein Rtc5 as a novel interactor of the V-ATPase. We show that Rtc5 localizes to the vacuole membrane depending on N-myristoylation and on its interactions with the V-ATPase. We further analyzed the influence of this protein, and the second yeast TLDc domain-containing protein, Oxr1, on V-ATPase function. We find that both Rtc5 and Oxr1 promote the disassembly of the vacuolar V-ATPase in vivo, counteracting the role of the assembly chaperone, the RAVE complex. Finally, Oxr1 is necessary for the retention in the late Golgi complex of an organelle-specific subunit of the V-ATPase. Collectively, our results shed light on the in vivo roles of yeast TLDc domain-containing proteins in relation to the V-ATPase, highlighting the multifaceted regulation of this crucial protein complex.

2024-03-11 | PXD046792 | Pride

Cross-link assisted spatial proteomics to map sub-organelle proteomes and membrane protein topology

Project description:The specific functions of cellular organelles and sub-compartments depend on their protein content, which can be characterized by spatial proteomics approaches. However, many spatial proteomics methods are limited in their ability to resolve organellar sub-compartments, profile multiple sub-compartments in parallel, and/or characterize membrane-associated proteomes. Here, we develop a cross-linking assisted spatial proteomics (CLASP) strategy that addresses these shortcomings. Using human mitochondria as a model system, we show that CLASP can elucidate spatial proteomes of all mitochondrial sub-compartments and provide topological insight into the mitochondrial membrane proteome in a single experiment. Biochemical and imaging-based follow-up studies demonstrate that CLASP allows discovering mitochondria-associated proteins and revising previous protein sub-compartment localization and membrane topology data. This study extends the scope of cross-linking mass spectrometry beyond protein structure and interaction analysis towards spatial proteomics, establishes a method for concomitant profiling of sub-organelle and membrane proteomes, and provides a resource for mitochondrial spatial biology

2024-02-21 | PXD046382 | Pride

Cross-link assisted spatial proteomics to map sub-organelle proteomes and membrane protein topology

2024-02-21 | PXD032132 | Pride

Don’t Waste Time – Ensure Success in Your Cross-Linking Mass Spectrometry Experiments Before You Begin.

Project description:Cross-linking mass spectrometry (XL-MS) is becoming a more popular tool for researchers to turn towards for studying proteins and their complexes of interest especially in complex samples such as lysates or whole-cells. Studying a targeted proteins in a complex mixture can be difficult as data on other higher abundant proteins may dominate, leaving little information on complexes of interest. Although it is known that cross-linking favours proteins of higher abundances, it is yet unclear what it means to be “abundant” to expect good cross-linking data. In this paper, we show that proteins of interest should be at least in the top 20 % abundance range to expect more than one cross-link found per protein. Furthermore, we show that a classical bottom-up LC-MS/MS experiment and iBAQ analysis can help determine the abundance level of proteins and a simple method to follow their enrichment if necessary. We hope that this guideline can be a starting point for researchers who would like to use XL-MS to study their protein of interest and help ensure a successful cross-linking experiment from the beginning.

2024-02-07 | PXD045792 | Pride

Conformation of the AP-3 complex explains its role in cargo recruitment at the Golgi

Project description:Crosslinking mass spectrometry study of an important adapter protein complex at the Golgi AP-3. We performed crosslinking mass spectrometry study using crosslinker DSSO and analyzed the digested peptides by LC-MS. The crosslinking study provided important information the subunit connectivity of AP-3, which facilitated the structural analysis of AP-3 by cyro-EM.

2022-02-17 | PXD028005 | Pride

Snapshot of in-cell protein contact sites reveals new host factors and hijacking of paraspeckles during influenza A virus infection

Project description:Influenza A virus (IAV) hijacks host cellular machinery, but many virus–IAV interactions and contacting protein sites remain uncharacterised, particularly those dependent on intact cellular architecture, such as membrane-associated or phase- separated compartments. Here, we applied in-cell cross-linking mass spectrometry (XL-MS), integrated with AlphaFold-based structural modelling and functional assays, to map protein-protein contact sites in IAV-infected human cells. This approach revealed previously unrecognised virus–host interactions linked to spatially organised processes, including the maturation pathway of HA through the membrane-bound ER– Golgi system, the novel interaction of M2 with the membrane-embedded LAT1 amino acid transporter, and the progressive disassembly of paraspeckles–phase-separated compartments in the nucleus. We validate M2-LAT1 interaction and paraspeckle disassembly in human primary lung epithelial cells and show that the paraspeckle disassembly constitutes a new and unique infection mechanism through which IAV releases RNA-binding proteins that support viral RNA replication. These findings advance the understanding of IAV manipulation of host cellular processes and illustrate how the integrative in-cell structural system biology approach captures native host-pathogen interactomes, infection pathways, and host cell perturbations.

2026-04-23 | PXD071226 | Pride

Hsp multichaperone complex buffers pathologically modified Tau

Project description:Alzheimer's disease is a neurodegenerative disorder in which misfolding and aggregation of pathologically modified Tau is critical for neuronal dysfunction and degeneration. The two central chaperones Hsp70 and Hsp90 coordinate protein homeostasis, but the nature of the interaction of Tau with the Hsp70/Hsp90 machinery has remained enigmatic. Here we show that Tau is a high-affinity substrate of the human Hsp70/Hsp90 machinery forming a 710 kDa client-loading complex. Complex formation involves extensive intermolecular contacts, blocks Tau aggregation and depends on Tau’s aggregation-prone repeat region. The Hsp90 co-chaperone p23 directly binds Tau and stabilizes the multichaperone/substrate complex, whereas the E3 ubiquitin-protein ligase CHIP efficiently disassembles the machinery targeting Tau to proteasomal degradation. Because phosphorylated Tau binds the Hsp70/Hsp90 machinery but is not recognized by Hsp90 alone, the data establish the Hsp70/Hsp90 multichaperone complex as a critical regulator of Tau in neurodegenerative disorders.

2022-08-11 | PXD032037 | Pride