Dataset Information

Global analysis of the human cytomegalovirus interactome identifies degradation hubs, domain associations and viral protein functions

ABSTRACT: Human cytomegalovirus (HCMV) is an important pathogen that extensively modulates host cells, downregulating >900 human proteins over the course of viral replication and degrading ≥133 proteins shortly after infection. The mechanism of degradation of most host proteins remains unresolved, and the functions of many viral proteins are incompletely characterised. We performed a systematic interactome analysis of 169 canonical HCMV proteins, and a subset of non-canonical HCMV proteins, in infected cells. This identified an extensive network of >3,400 virus-host and >150 virus-virus protein interactions, providing insights into novel functions for multiple viral genes. Domain analysis predicted binding of the viral UL25 protein to the SH3 domains of NCK Adaptor Protein 1. Viral interacting proteins were identified for 31/133 degraded host targets. Finally, the uncharacterised, non-canonical ORFL147C protein was found to interact with elements of the mRNA splicing machinery, and a mutational study suggested its importance in viral replication. The interactome data will be important for future studies of herpesvirus infection.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human) Human Herpesvirus 5 Strain Merlin Betaherpesvirinae

TISSUE(S): Cell Culture, Fibroblast

SUBMITTER: Michael Weekes

LAB HEAD: Michael Weekes

PROVIDER: PXD014845 | Pride | 2020-01-22

REPOSITORIES: Pride

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Publications

Human cytomegalovirus interactome analysis identifies degradation hubs, domain associations and viral protein functions.

Nobre Luis V LV Nightingale Katie K Ravenhill Benjamin J BJ Antrobus Robin R Soday Lior L Nichols Jenna J Davies James A JA Seirafian Sepehr S Wang Eddie Cy EC Davison Andrew J AJ Wilkinson Gavin Wg GW Stanton Richard J RJ Huttlin Edward L EL Weekes Michael P MP

eLife 20191224

Human cytomegalovirus (HCMV) extensively modulates host cells, downregulating >900 human proteins during viral replication and degrading ≥133 proteins shortly after infection. The mechanism of degradation of most host proteins remains unresolved, and the functions of many viral proteins are incompletely characterised. We performed a mass spectrometry-based interactome analysis of 169 tagged, stably-expressed canonical strain Merlin HCMV proteins, and two non-canonical HCMV proteins, in infected ...[more]

PMID: 31873071

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Project description:We have established that human cytomegalovirus (HCMV) infection modulates the biology of target primary blood monocytes, allowing HCMV to use monocytes as 'vehicles' for its systemic spread. HCMV infection of monocytes results in rapid induction of PI(3)K and NF-kB activity. Integrins, which are upstream of the PI(3)K and NF-kB pathways, were shown to be involved in HCMV binding to and entry into fibroblasts, suggesting that receptor-ligand-mediated signaling following viral binding to integrins on monocytes could trigger the functional changes seen in infected monocytes. We now show that integrin engagement and the activation of the integrin/Src-signaling pathway is essential for the induction of HCMV-infected monocyte motility. To investigate how integrin engagement by HCMV triggers monocyte motility, we examined the infected monocyte transcriptome and found that the integrin/Src-signaling pathway regulates the expression of paxillin, which is an important signal transducer in the regulation of actin rearrangement during cell adhesion and movement. Functionally, we observed that paxillin is activated via the integrin/Src-signaling pathway and is required for monocyte motility. Because motility is intimately connected to cellular cytoskeletal organization, a process that is also important in viral entry, we investigated the role paxillin regulation plays in the process of viral entry of monocytes. New results confirmed that HCMV`s ability to enter target monocytes is significantly inhibited in cells deficient in paxillin expression or that had their integrin/Src/paxillin signaling pathway blocked. From our data, HCMV-cell interactions emerge as an essential trigger for the cellular changes that allow for HCMV entry and hematogenous dissemination. Monocytes were mock-infected, HCMV-infected, or pretreated with PP2 inhibitor prior to HCMV infection. There were three samples analyzed per individual replicate. Three replicates are included. comparative studies with a use of the specific Src kinase activity inhibitor

Dataset Information

Global analysis of the human cytomegalovirus interactome identifies degradation hubs, domain associations and viral protein functions

Publications

Human cytomegalovirus interactome analysis identifies degradation hubs, domain associations and viral protein functions.

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