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Proteome characterization of HCMV virus microenvironment

ABSTRACT: The control of host processes which influence cell-cell communication is central to determining the outcome of a virus infection in tissue. Despite this, it remains unclear how cells either in close or distant proximity to an infected cell differentially respond to the primary infection. Here, we established a virus infection microenvironment to resolve molecular features and functional consequences of cell spatial address within this localized niche by mass spectrometry.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human) Human Betaherpesvirus 5

TISSUE(S): Cell Culture, Fibroblast

SUBMITTER: Bokai Song

LAB HEAD: Ileana Cristea

PROVIDER: PXD038547 | Pride | 2023-05-11

REPOSITORIES: Pride

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			Action	DRS
	20200905_G-M-_rep1.msf	Msf
	20200905_G-M-_rep1.raw	Raw
	20200905_G-M-_rep2.msf	Msf
	20200905_G-M-_rep2.raw	Raw
	20200905_G-M-_rep3.msf	Msf

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Publications

Intercellular communication within the virus microenvironment affects the susceptibility of cells to secondary viral infections.

Song Bokai B Sheng Xinlei X Justice Joshua L JL Lum Krystal K KK Metzger Peter J PJ Cook Katelyn C KC Kostas James C JC Cristea Ileana M IM

Science advances 20230510 19

Communication between infected cells and cells in the surrounding tissue is a determinant of viral spread. However, it remains unclear how cells in close or distant proximity to an infected cell respond to primary or secondary infections. We establish a cell-based system to characterize a virus microenvironment, distinguishing infected, neighboring, and distal cells. Cell sorting, microscopy, proteomics, and cell cycle assays allow resolving cellular features and functional consequences of proxi ...[more]

PMID: 37163594

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Proteome characterization of HCMV virus microenvironment with addition of cidofovir

Project description:The control of host processes which influence cell-cell communication is central to determining the outcome of a virus infection in tissue. Despite this, it remains unclear how cells either in close or distant proximity to an infected cell differentially respond to the primary infection. Here, we established a virus infection microenvironment to resolve molecular features and functional consequences of cell spatial address within this localized niche by mass spectrometry. To rule out the possibility that neighboring cells are infected by the viral progeny from the primary infection, cidofovir is added in the system to prevent viral genome replication in order to prevent the spread of infection.

2023-05-11 | PXD040582 | Pride

Comparative quantitative proteomic analysis of HCMV infected cells highlights pUL138 as a multiple functional protein

Project description:Human cytomegalovirus (HCMV) is a widespread virus and can establish life-long latent infection in large populations. In order to establish persistent and latent infection in healthy individuals, HCMV encodes a large array of proteins that can modulate different components and pathways of host cells. It has been reported that pUL138 encoded by UL133-UL138 polycistronic locus promotes a latent infection in primary CD34+ HPCs infected in vitro. In this study, a recombinant HCMV, namely HanUL138del, was constructed by deleting the UL138 locus of Han, a clinic HCMV strain. Then a comparative quantitative proteomic analysis of Han and HanUL138del infected MRC5 cells was performed, aiming to study the effect of pUL138 on host cell in the context of HCMV infection. Our result indicated that at the early phase of HCMV infection, innate immune response was differentially activated, while at late phase of HCMV infection, multiple host proteins were differentially expressed, between Han or HanUL138del infected cells.

2021-09-08 | PXD015931 | Pride

SIRT3 interactions and mitochondrial proteome during HCMV infection

Project description:SIRT3 has been shown to inhibit HCMV infection, but the mechanism under the antiviral effect is not clear. To identify the mediator of SIRT3 antiviral function, we aim to identify and quantify SIRT3 interactions during HCMV infection. A set of large-scale IPs were performed to determine the specific interactions with SIRT3 during infection, while small-scale IPs were conducted to determine the temporal interactions over the life cycle of infection. The mitochondrial proteome was used to characterize the changes of protein abundances after infection, and it was used for the normalization of acetylation.

2021-04-01 | PXD018995 | Pride

Effect of isoflavone agonists of IRF-3 on response to RNA viruses

Project description:There is a growing need for novel antiviral therapies that are broad-spectrum, effective, and not subject to resistance due to viral mutations. Using high-throughput screening methods, including computational docking studies and an ISG54-luciferase reporter assay, we identified a class of isoflavone compounds that act as specific agonists of innate immune signaling pathways and cause activation of the IRF-3 transcription factor. The objective of the microarray study was to examine the biological pathways associated with global gene expression changes following agonist treatment. Total RNA isolation and mRNA amplification were performed on equal masses of total RNA from MRC5 cells treated with either DMSO (negative control; n=3), or 10μM of the isoflavone agonist KIN 101 (n=3) at 20 hours post treatment. As a positive control for response to an RNA virus, total RNA isolation and mRNA amplification was performed on equal masses of total RNA from MRC5 cells infected with Sendai virus (n=3) at 20 hours post infection.

2012-05-17 | E-GEOD-38016 | biostudies-arrayexpress

Identification of interactors with HCMV pTRS1 using BioID and proteomics

Project description:Human cytomegalovirus (HCMV) is a ubiquitous pathogen that infects the majority of the world’s population. Lytic HCMV replication in immunocompromised individuals or neonates can lead to severe disease in multiple organ systems and even death. The establishment of lytic replication is driven by the first viral proteins expressed upon infection, the immediate early proteins, which play a key role in creating an intracellular environment conducive to virus replication. Two immediate early proteins, the functional orthologs pTRS1 and pIRS1, stimulate immediate early gene expression by suppressing antiviral PKR/eIF2a signaling and enhance the translation of viral mRNAs independent of PKR antagonism. To better understand the molecular functions of pTRS1, we used proximity labeling proteomics to identify proteins that interact with pTRS1 in infected cells. Multiple novel host and viral interactors were identified, including the catalytic subunits of the protein phosphatase 1 (PP1) holoenzyme. Mutations to a PP1 catalytic subunit known to disrupt binding to PP1 regulatory subunits decreased binding to pTRS1. pTRS1 immune complexes contained phosphatase activity, and inhibition of phosphatase activity in transfected or infected cells reversed the ability of pTRS1 to inhibit the antiviral kinase PKR. Depletion of individual PP1 catalytic subunits decreased virus replication and increased the phosphorylation of the PKR substrate eIF2a. Together our data suggest potential novel functions for pTRS1, and define a novel role for PP1 as an antagonist of the antiviral PKR/eIF2 signaling axis during HCMV infection.

2024-10-11 | PXD053924 | Pride

RNA-Seq of influenza A/Puerto Rico/8/1934 (H1N1) virus infected MRC5, WI-38 VA-13, A549 and HEK293FT human cells

Project description:To study genes involved in cellular response to Influenza A virus infection, human cells MRC5, WI-38 VA-13, A549 and HEK293FT were infected with influenza A/Puerto Rico/8/1934 (H1N1) virus. RNA-seq analysis of infected cell lines after 48 h was performed on an Illumina NextSeq 500 platform. The same mock-infected cells were used as controls. PolyA RNA-enriched fraction was used for constructing of cDNA libraries. Differential expressed genes were identified using R package DESeq2.

2020-11-23 | E-MTAB-9511 | biostudies-arrayexpress

Club cells surviving influenza A virus infection induce temporary non-specific anti-viral immunity

Project description:Cells respond differently to influenza virus infection after already having been infected previously. Anlysis of cellular RNA after influenza virus infection

2016-03-01 | E-GEOD-77734 | biostudies-arrayexpress

Temporal dynamics of protein complex formation and dissociation during human cytomegalovirus infection

Project description:Emerging evidence indicates that viral infection alters protein complex composition to induce both proviral and antiviral effects. However, global changes in protein complexes during infection remains understudied. Here, we employ thermal proximity coaggregation (TPCA) analysis to provide the first global and temporal characterization of changes in protein complexes during human cytomegalovirus (HCMV) infection. This dataset provides a resource to mine the alterations in protein complexes during HCMV infection. Furthermore, our subsequent experiments identified that these changes in protein complex composition and interactions functionally contribute to the progression of infection.

2020-01-29 | PXD014747 | Pride

T cell UTX promotes Tfh-mediated clearance of persistent virus infection

Project description:Epigenetic changes, including histone methylation, control T cell differentiation and memory formation, though the enzymes that mediate these processes are not clear. We show that UTX, a histone H3 lysine 27 (H3K27) demethylase, promotes the generation of T follicular helper (Tfh) cells, a CD4+ T cell subset essential for B cell antibody generation and clearance of chronic viral infections. Mice with T cell specific UTX deletion (UTX TKO mice) had fewer Tfh cells, showed defective germinal center formation, lacked virus-specific IgG production, and were unable to resolve chronic lymphocytic choriomeningitis virus infection. In UTX TKO T cells, decreased expression of IL-6R±, and other Tfh-related loci, was associated with increased H3K27 methylation. Additionally, Turner Syndrome subjects, who are predisposed to chronic ear infections, have reduced UTX expression in immune cells and decreased Tfh frequency. Thus, we identify a critical link between UTX in T cells and immunity to infection. Mice were infected with chronic LCMV and sorted for Tfh on day 21 pi (sorted for: CD4+CXCR5+CD62Llo T cell). ChIP-Seq and RNA-Seq on WT and Utx KO cells.

2015-09-01 | E-GEOD-64969 | biostudies-arrayexpress

Transcription profiling of human EBV infected EBNA-3 positive and negative human B lymphocytes

Project description:The genome of Epstein-Barr virus (EBV) encodes 86 proteins but only a limited set is expressed in EBV-growth transformed B cells, termed lymphoblastoid cell lines (LCLs). These cells proliferate via the concerted action of EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), some of which are rate limiting to establish a stable homeostasis of growth promoting and anti-apoptotic activities. We show here that EBV mutants, which lack the EBNA-3A gene, are impaired but can still initiate cell-cycle entry and proliferation of primary human B cells in contrast to an EBNA-2-deficient mutant virus. Surprisingly and in contrast to previous reports, these viral mutants are attenuated in growth transformation assays but give rise to permanently growing EBNA-3A negative B cell lines which exhibit reduced proliferation rates and elevated levels of apoptosis. Expression profiles of EBNA-3A deficient LCLs are characterized by 129 upregulated and 167 downregulated genes, which are significantly enriched for genes involved in apoptotic processes or cell cycle progression like the tumor suppressor gene p16/INK4A or might contribute to essential steps in the viral life cycle. In addition EBNA-3A cellular target genes remarkably overlap with previously identified targets of EBNA-2. Experiment Overall Design: Comparison of gene expression profiles from lymphoblastoid cell lines (LCLs) infected with EBNA-3A positive or negative EBV. LCLs were generated from three donors. A total of five EBNA-3A positive and nine negative LCLs were analysed.

2009-09-13 | E-GEOD-17908 | biostudies-arrayexpress

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