Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Quantitative comparative analysis of human erythrocyte surface proteins between individuals from two genetically distinct populations

ABSTRACT: Red blood cells (RBC) play a critical role in oxygen transport, and are the focus of important human diseases including malaria and the haemoglobinopathies. Although previous studies aimed at classifying the RBC proteome have examined whole or membrane-enriched fractions from RBC, none have used specific strategies directed at enriching proteins with exposed extracellular domains. Furthermore, there has been no systematic analysis of variation in abundance of RBC surface proteins either within or between genetically disparate human populations. These questions are of particular importance to inform not only basic RBC biology but additionally to identify novel candidate Plasmodium receptors, where selective pressures have moulded the RBC surface, exemplified by genetic absence of Duffy antigen in populations from sub-Saharan Africa. Here, we use ‘Plasma membrane profiling’ and tandem mass tag-based mass spectrometry to specifically enrich and quantify cell surface proteins from primary RBC from two sets of nine donors from the UK or Senegal. We define a ‘stringent’ and ‘sensitive’ RBC surface proteome and identify potential candidate Plasmodium receptors on the basis either of diminished protein abundance, or increased variation in West African individuals compared to people from the UK.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Erythrocyte, Blood

SUBMITTER: Michael Weekes

LAB HEAD: Michael Weekes

PROVIDER: PXD013852 | Pride | 2019-08-22

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	PRIDEsubmissionRavenhilletal.docx	Other
	RBCPeptides.xlsx	Xlsx
	RBCProteins.xlsx	Xlsx
	Senegaldonorsfraction1.raw	Raw
	Senegaldonorsfraction2.raw	Raw

Items per page:

1 - 5 of 17

Similar Datasets

Comprehensive cell surface proteomics defines markers of classical, intermediate and non-classical monocytes

Project description:Monocytes are a critical component of the cellular innate immune system, and can be subdivided into classical, intermediate and non-classical subsets on the basis of surface CD14 and CD16 expression. Classical monocytes play the canonical role of phagocytosis, and account for the majority of circulating cells. Intermediate and non-classical cells are known to exhibit varying levels of phagocytosis and cytokine secretion, and are differentially expanded in certain pathological states. Characterisation of cell surface proteins expressed by each subset is informative not only to improve understanding of phenotype, but also to provide biological insight into function. Here we use highly multiplexed Tandem-Mass-Tag (TMT)-based mass spectrometry with selective cell surface biotinylation to characterise the classical monocyte surface proteome, then interrogate the phenotypic differences between each monocyte subset to identify novel protein markers.

2020-03-03 | PXD013832 | Pride

Comparative cell surface proteomic analysis of the primary human T cell and monocyte responses to type I interferon

Project description:The cellular response to interferon (IFN) is essential for antiviral immunity, IFN-based therapy and IFN-related disease. The plasma membrane (PM) provides a critical interface between the cell and its environment, and is the initial portal of entry for viruses. Nonetheless, the effect of IFN on PM proteins is surprisingly poorly understood, and has not been systematically investigated in primary immune cells. Here, we use multiplexed proteomics to quantify IFNα-stimulated PM protein changes in primary human CD14+ monocytes and CD4+ T cells from five donors, quantifying 606 and 482 PM proteins respectively. Comparison of cell surface proteomes revealed a remarkable invariance between donors in the overall composition of the cell surface from each cell type, but a marked donor-to-donor variability in the effects of IFNα. Furthermore, whereas only 2.7% of quantified proteins were consistently upregulated by IFNα at the surface of CD4+ T cells, 6.8% of proteins were consistently upregulated in primary monocytes, suggesting that the magnitude of the IFNα response varies according to cell type. Amongst these differentially regulated proteins, we found the viral target Endothelin-converting enzyme 1 (ECE1) to be an IFNα-stimulated protein exclusively upregulated at the surface of CD4+ T cells. We therefore provide a comprehensive map of the cell surface of IFNα-stimulated primary human immune cells, including previously uncharacterised interferon stimulated genes (ISGs) and candidate antiviral factors.

2021-02-09 | PXD022834 | Pride

Temporal proteomic analysis of BK polyomavirus infection reveals virus-induced G2 arrest and highly effective evasion of innate immune sensing

Project description:BK polyomavirus (BKPyV) is a small DNA virus that establishes a life-long persistent infection in the urinary tract of most people. BKPyV is known to cause severe morbidity in renal transplant recipients and can lead to graft rejection. The simple 5.2 kilobase pair dsDNA genome expresses just seven known proteins, thus it relies heavily on host machinery to replicate. How the host proteome changes over the course of infection is key to understanding this host:virus interplay. Here for the first time quantitative temporal viromics has been used to quantify global changes in >9,000 host proteins in two types of primary human epithelial cell throughout 72 hours of BKPyV infection. These data demonstrate the importance both of cell cycle progression and pseudo-G2 arrest in effective BKPyV replication, along with a surprising lack of innate immune response throughout the whole virus replication cycle. BKPyV thus evades pathogen recognition to prevent activation of innate immune responses in a sophisticated manner.

2019-05-23 | PXD013940 | Pride

Quantitative temporal proteomic analysis of vaccinia virus infection reveals regulation of histone deacetylases by an interferon antagonist

Project description:Vaccinia virus (VACV) has numerous immune evasion strategies, including multiple mechanisms of inhibition of IRF-3, NF-κB and type I interferon (IFN) signaling. Here, we used highly multiplexed proteomics to quantify >8,000 cellular proteins and ~80% of viral proteins over seven time points spanning the whole course of VACV infection. This identified multiple novel viral targets, including putative natural killer cell ligands and IFN-stimulated genes. The class II histone deacetylase HDAC5 was selectively degraded early during VACV infection. Use of cell lines in which HDAC5 was overexpressed or knocked out showed that HDAC5 restricted replication of both VACV and herpes simplex virus type 1 (HSV-1). By generating a protein-based temporal classification of VACV gene expression, we identified the early protein C6, a multifunctional IFN antagonist, as the factor that targets HDAC5 for proteasomal degradation. Our approach thus identifies both a novel restriction factor and a viral mechanism of innate immune evasion.

2019-05-07 | PXD012785 | Pride

HCMV US2 co-opts TRC8 to degrade the ER-resident protein LMAN2L

Project description:The human cytomegalovirus (HCMV) pUS2 glycoprotein exploits the host’s endoplasmic reticulum (ER)-associated degradation (ERAD) pathway to degrade MHC class I and prevent antigen presentation. Beyond MHC class I, pUS2 has been shown to target a range of cellular proteins for degradation, preventing their cell surface expression. Here we have identified a novel pUS2 target, ER-resident protein lectin mannose binding 2 like (LMAN2L). pUS2 expression was both necessary and sufficient for the downregulation of LMAN2L, which was dependent on the cellular E3 ligase TRC8. Given the hypothesised role of LMAN2L in the trafficking of glycoproteins, we employed proteomic plasma membrane profiling to measure LMAN2L-dependent changes at the cell surface. Known pUS2 target integrin alpha-6 (ITGA6) was downregulated from the surface of LMAN2L deficient cells. Overall, these results suggest a novelstrategy of pUS2-mediated protein degradation whereby pUS2 targets LMAN2L to impair trafficking of ITGA6. Given US2 can directly target other integrins, this single viral protein exhibits both direct and indirect mechanisms to downregulate integrins from the cell surface.

2024-06-23 | PXD050878 | Pride

ADAM17 targeting by human cytomegalovirus remodels the cell surface proteome to simultaneously regulate multiple immune pathways

Project description:Human cytomegalovirus (HCMV) is a major human pathogen whose life-long persistence is enabled by its remarkable capacity to systematically subvert host immune defences. In exploring the finding that HCMV infection upregulates tumor necrosis factor receptor 2 (TNFR2), a ligand for the proinflammatory anti-viral cytokine TNFa, we discovered the underlying mechanism was due to targeting of the protease, A Disintegrin And Metalloproteinase 17 (ADAM17). ADAM17 is the prototype ‘sheddase, a family of proteases that cleaves other membrane-bound proteins to release biologically active ectodomains into the supernatant. HCMV impaired ADAM17 surface expression through the action of two virally-encoded proteins in its UL/b’ region, UL148 and UL148D. Proteomic plasma membrane profiling of cells infected with a HCMV double deletion mutant for UL148 and UL148D with restored ADAM17 expression, combined with ADAM17 functional blockade, showed that HCMV stabilized the surface expression of 114 proteins (p<0.05) in an ADAM17-dependent fashion. These included known substrates of ADAM17 with established immunological functions such as TNFR2 and Jagged1, but also numerous novel host and viral targets, such as Nectin1, UL8 and UL144. Regulation of TNFa-induced cytokine responses and NK inhibition during HCMV infection were dependent on this impairment of ADAM17. We therefore identify a viral immunoregulatory mechanism in which targeting a single sheddase enables broad regulation of multiple critical surface receptors, revealing a paradigm for viral-encoded immunomodulation.

2023-10-11 | PXD044082 | Pride

Proteomic analysis of circulating immune cells identifies novel cellular phenotypes associated with COVID-19 severity

Project description:Certain serum proteins, including CRP and D-dimer, have prognostic value in patients with SARS-CoV-2 infection. Nonetheless, these factors are non-specific, and provide limited mechanistic insight into the peripheral blood mononuclear cell (PBMC) populations which drive the pathogenesis of severe COVID-19. To identify novel cellular phenotypes associated with disease progression, we here describe a comprehensive, unbiased analysis of the total and plasma membrane proteomes of PBMCs from a cohort of 40 unvaccinated individuals with SARS-CoV-2 infection, spanning the whole spectrum of disease severity. Combined with RNA-seq and flow cytometry data from the same donors, we define a comprehensive multi-omic profile for each severity level, revealing cumulative immune cell dysregulation in progressive disease. In particular, the cell surface proteins CEACAMs1, 6 and 8, CD177, CD63 and CD89 are strongly associated with severe COVID-19, corresponding to the emergence of atypical CD3+CD4+CD177+ and CD16+CEACAM1/6/8+ mononuclear cells. Utilisation of these markers may facilitate real-time patient assessment by flow cytometry, and identify immune cell populations that could be targeted to ameliorate immunopathology.

2023-03-29 | PXD040703 | Pride

Human cytomegalovirus protein pUL36: a dual cell death pathway inhibitor

Project description:Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of intrinsic, innate and adaptive viral immune evasion. Here, we employ multiplexed tandem mass tag-based proteomics to characterise host proteins targeted for degradation late during HCMV infection. This approach revealed that mixed lineage kinase domain-like protein (MLKL), a key terminal mediator of cellular necroptosis, was rapidly and persistently degraded by the minimally passaged HCMV strain Merlin but not the extensively passaged strain AD169. The strain Merlin viral inhibitor of apoptosis pUL36 was necessary and sufficient both to degrade MLKL and to inhibit necroptosis. Furthermore, mutation of pUL36 Cys131 abrogated MLKL degradation and restored necroptosis. As the same residue is also required for pUL36-mediated inhibition of apoptosis by preventing proteolytic activation of pro-caspase 8, we define pUL36 as a multifunctional inhibitor of both apoptotic and necroptotic cell death.

2020-07-10 | PXD017279 | Pride

Human cytomegalovirus protein RL1 degrades the antiviral factor SLFN11 via recruitment of the CRL4 E3 ubiquitin ligase complex

Project description:Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of viral immune evasion, targeting intrinsic, innate and adaptive immunity. We have employed two novel, orthogonal multiplexed tandem mass tag-based proteomic screens to identify host proteins downregulated by viral factors expressed during the latest phases of viral infection. This approach revealed that the HIV-1 restriction factor Schlafen-11 (SLFN11) was degraded by the poorly characterised, late-expressed HCMV protein RL1, via recruitment of the Cullin4-RING E3 Ubiquitin Ligase (CRL4) complex. SLFN11 potently restricted HCMV infection, inhibiting the formation and spread of viral plaques. Overall, we show that a restriction factor previously thought only to inhibit RNA viruses additionally restricts HCMV. We define the mechanism of viral antagonism and also describe an important resource for revealing additional molecules of importance in antiviral innate immunity and viral immune evasion.

2022-02-22 | PXD026785 | Pride

Rapid degradation pathways of host proteins during HCMV infection revealed by quantitative proteomics

Project description:Human cytomegalovirus is an important pathogen in immunocompromised individuals and neonates, and a paradigm for viral immune evasion. We previously developed a quantitative proteomic approach that identified 133 proteins degraded during the early phase of HCMV infection, including known and novel antiviral factors. The majority were rescued from degradation by MG132, which is known to inhibit lysosomal cathepsins in addition to the proteasome. Global definition of the precise mechanisms of host protein degradation is important both to improve our understanding of viral biology, and to inform novel antiviral therapeutic strategies. We therefore developed and optimised a multiplexed comparative proteomic analysis using the selective proteasome inhibitor bortezomib in addition to MG132, to provide a global mechanistic view of protein degradation. Of proteins rescued from degradation by MG132, 62-85% were also rescued by bortezomib, suggesting both that the predominant mechanism of protein degradation employed by HCMV is via the proteasome, and that alternative pathways for degradation are nevertheless important. Our approach and data will enable improved mechanistic understanding of HCMV and other viruses, and provide a shortlist of candidate restriction factors for further analysis.

2021-01-08 | PXD021961 | Pride