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AP-MS analysis of m7GTP-binding proteins in Murine norovirus infected BV-2 cells (v2)

ABSTRACT: SILAC-labelling analysis of murine BV-2 cells infected at high multiplicity of infection (10 TCID50/cell) and harvested at 4h or 9h post-infection. Lysates were then suject to m7GTP-sepharose affinity purification to enrich for translation initiation factors. As norovirus translation uses a VPg protein covalently linked to the 5' of its RNAs for initiation factor recruitment it was hypothesised that norovirus infection could modify initiation factor complexes. A paired dataset investigates the effects of MNV infection on total protein levels in infected cells. This dataset is a Maxquant reanalysis of data previously submitted under PXD004019

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Dendritic Cell, Cell Culture

SUBMITTER: Edward Emmott

LAB HEAD: Ian Goodfellow

PROVIDER: PXD004983 | Pride | 2017-01-17

REPOSITORIES: Pride

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			Action	DRS
	BV2_1.raw	Raw
	BV2_2.raw	Raw
	BV2_3.raw	Raw
	Howtoviewspectra.txt	Txt
	fasta.zip	Other

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Publications

Norovirus-Mediated Modification of the Translational Landscape via Virus and Host-Induced Cleavage of Translation Initiation Factors.

Emmott Edward E Sorgeloos Frederic F Caddy Sarah L SL Vashist Surender S Sosnovtsev Stanislav S Lloyd Richard R Heesom Kate K Locker Nicolas N Goodfellow Ian I

Molecular & cellular proteomics : MCP 20170113 4 suppl 1

Noroviruses produce viral RNAs lacking a 5' cap structure and instead use a virus-encoded viral protein genome-linked (VPg) protein covalently linked to viral RNA to interact with translation initiation factors and drive viral protein synthesis. Norovirus infection results in the induction of the innate response leading to interferon stimulated gene (ISG) transcription. However, the translation of the induced ISG mRNAs is suppressed. A SILAC-based mass spectrometry approach was employed to analy ...[more]

PMID: 28087593

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AP-MS analysis of m7GTP-binding proteins in Murine norovirus infected BV-2 cells

Project description:SILAC-labelling analysis of murine BV-2 cells infected at high multiplicity of infection (10 TCID50/cell) and harvested at 4h or 9h post-infection. Lysates were then suject to m7GTP-sepharose affinity purification to enrich for translation initiation factors. As norovirus translation uses a VPg protein covalently linked to the 5' of its RNAs for initiation factor recruitment it was hypothesised that norovirus infection could modify initiation factor complexes. A paired dataset investigates the effects of MNV infection on total protein levels in infected cells (PXD004015).

2016-07-07 | PXD004019 | Pride

Quantitative proteomic analysis of Murine Norovirus infected BV-2 cells

Project description:SILAC-labelling analysis of murine BV-2 cells infected at high multiplicity of infection (10 TCID50/cell) and harvested at 4h or 9h post-infection. Represents the first quantitative proteomics analysis of norovirus-infected cells. A paired AP-MS dataset investigates the effects of MNV infection on eukaryotic initiation factor (eIF) complex formation with this dataset providing data on the overall abundance of eIF components in infected cells.

2016-06-28 | PXD004015 | Pride

Quantitative proteomic analysis of Murine Norovirus infected BV-2 cells (v2)

Project description:SILAC-labelling analysis of murine BV-2 cells infected at high multiplicity of infection (10 TCID50/cell) and harvested at 4h or 9h post-infection. Represents the first quantitative proteomics analysis of norovirus-infected cells. A paired AP-MS dataset investigates the effects of MNV infection on eukaryotic initiation factor (eIF) complex formation with this dataset providing data on the overall abundance of eIF components in infected cells. This is a reanalysis of an earlier dataset (PXD004015) performed in Maxquant (v1.5.5.1).

2017-01-17 | PXD004984 | Pride

Phosphoproteomic analysis of Bluetongue virus infection

Project description:Bluetongue virus causes infections in wild and domesticated ruminants, with the potential for causing significant morbidity, mortality and economic harm in both the developing and developed world. While vaccines have been developed, no treatment for acute infections exists. In this regard, a possible approach is to determine host-cell factors utilised by Bluetongue virus. We thus proceeded to characterize the phosphoproteome of BTV infected HeLa cells to elucidate the intracellular signalling pathways and identify critical host factors activated during Bluetongue virus infection.

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Identification of hypoxia-induced proteins using SILAC-based proteomics

Project description:A hallmark of solid tumours is the development of hypoxic regions where rapidly growing transformed cells outstrip their blood supply. Tumour cells in these starved regions sense reduced levels of oxygen and switch on genes that help them to adapt, survive and continue growing. Since hypoxia is a key physiological difference between normal and cancer tissue, an opportunity exists to selectively kill tumour cells by exploiting the differences in protein expression between normal and hypoxic tumour tissue. However, a major challenge is to identify druggable hypoxia-induced proteins critical for tumour cell survival. This is especially important in cancers of unmet need where hypoxia gene signatures correlate with poor prognosis (for example, colorectal cancers). To identify new hypoxia-induced proteins, we performed SILAC-based proteomics on colorectal cancer cells exposed to normoxia and hypoxia. In this study, we identify a new hypoxia-activated GPCR signalling axis that enables colorectal tumour cells to survive the microenvironmental stress of hypoxia. Our findings uncover a previously unappreciated role for this GPCR-axis as a key regulator of the adaptive response to hypoxia and highlght an opportunity to exploit tumour-associated hypoxia for therapy.

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A novel paracrine induction of RNA Granules by Feline Calicivirus

Project description:We are here presenting a new paracrine induction of RNA granules by viruses. Infection by viruses imposes major stress on the host cell. In response to this stress, infected cells can induce several defence mechanisms, which include the activation of stress response pathways and the innate immune response. These often result in an inhibition of translation culminating in the assembly of cytoplasmic granules called stress granules (SGs). SGs assembly follows from liquid phase separation of aggregation-prone proteins such G3BP1 and TIA-1, leading to the sequestration of mRNAs. Because this threatens viral gene expression, viruses need to evade these stress response pathways to propagate. Using feline calicivirus (FCV), surrogate for norovirus, the main virus responsible for gastroenteritis outbreaks worldwide, we previously showed that FCV impairs SGs assembly by cleaving the scaffold protein G3BP1. Interestingly, we observed that uninfected bystander cells assembled G3BP1 granules, suggesting a paracrine response trigged by the infection. We now present evidence that virus-free supernatant generated from infected cells can induce the formation of RNA granules. We have characterised the dynamic of the granules assembly via confocal microscopy. Moreover, we provide an understanding of paracrine granules function and specificity through their affinity purification followed by proteomics and RNAseq analysis of their proteins and mRNAs content. This helps to define rules of assembly and novel functions for paracrine granules highlighting fundamental differences with canonical stress granules.

2022-03-14 | PXD021881 | Pride

AP-MS analysis of murine and human norovirus VPg interactomes

Project description:The norovirus VPg protein is covalently linked to the viral genome in place of a 5' cap, and functions as a cap-substitute, capable of interacting with translation initiation factors. Following on from our previous study (Chung et. al. 2014, J. BIol. Chem.) we wished to determine the interactome of human norovirus VPg, and compare that of murine norovirus VPg. We had previously demonstrated that mutation of the penultimate C-terminal phenylalanine residue in murine norovirus VPg greatly reduced initiation factor binding (F123A). Insertion of the equivalent mutation into human norovirus (F137A) also reduced initiation factor binding. Affinity purification of wild-type of mutant human and murine norovirus VPg was accomplished using GFP-tagged VPg transfected into SILAC-labelled human HEK-293T cells.

2019-09-09 | PXD007585 | Pride

Mass spectrometry of immunoprecipitations from control and Er blood group

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2024-01-26 | PXD035013 | Pride

Human Platelet PIP3 Signalosome

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2017-06-06 | PXD003777 | Pride

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