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

Ravenhill, Benjamin J.; Kanjee, Usheer; Ahouidi, Ambroise D.; Nobre, Luis; Williamson, James C; Goldberg, Jonathan M.; Antrobus, Robin; Diéye, Tandakha Ndiaye; Duraisingh, Manoj T.; Weekes, Michael P.

doi:10.1038/s42003-019-0596-y

Cited by 25 publications

(23 citation statements)

References 61 publications

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“…C -Classical monocyte sample, I -Intermediate monocyte sample, NC -Non-classical monocyte sample. (2020) 10:4560 | https://doi.org/10.1038/s41598-020-61356-w www.nature.com/scientificreports www.nature.com/scientificreports/ other cell surface proteomic analyses in primary cells have been limited to CD4+ T-cells 16 , erythrocytes 17,18 , NK cells, adipocytes and a subset of tumour cells 19 . We quantify 373 cell surface proteins from classical monocytes, and further quantify 313 proteins across all monocyte subsets.…”

Section: Discussionmentioning

confidence: 99%

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

Ravenhill

Soday

Houghton

et al. 2020

Sci Rep

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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 may also provide biological insights 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. Monocytes play critical roles in the response to infection and inflammation and antigen presentation. Key effector functions are mediated by differentiated cells, including macrophages and myeloid dendritic cells. Monocytes can be divided into subsets based on the presence of the cell surface lipopolysaccharide (LPS) co-receptor CD14, and Fc-Gamma Receptor III (CD16). CD14++ CD16− classical monocytes account for ~85% of circulating monocytes, intermediate cells (CD14++ CD16+) for ~5% and non-classical cells (CD14+ CD16++) for ~10% of circulating monocytes 1,2. It has become increasingly well established that each subset plays divergent roles in different diseases, as well as differing in the ability to secrete cytokines and respond to pathogen associated molecular patterns (PAMPs). Classical monocytes are phagocytic and readily secrete inflammatory cytokines. Conversely, the CD16 positive non-classical cells are poorly phagocytic and are suggested to secrete TNFα in response to some stimuli, but less of other pro-inflammatory molecules 2-4. Intermediate monocytes are increased in diseases such as severe asthma, rheumatoid arthritis and sarcoidosis, and there is some evidence for expansion of classical monocytes in atherosclerosis 5-8. It is still unclear whether intermediate monocytes represent a truly distinct monocyte subset, or merely a transitional stage between classical and non-classical cells 9. Cells of the innate and adaptive immune systems can be categorised on the basis of microscopic appearance and expression of plasma membrane (PM) proteins, enabling separation by fluorescence activated cell sorting (FACS). As such, systematic evaluation of the entire cell surface proteome expressed by a given immune cell population is a powerful tool to characterise cellular function and distinguish cell types. Previous studies of monocytes subsets have examined transcriptional differences and offer varying depths of information about subset markers, suitability of individual proteins for discriminating subsets by cell surface flow cytometry and the importance of each protein...

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Section: Discussionmentioning

confidence: 99%

“…Data analysis. Data analysis was performed with modifications as previously described 18,38 . In the following description, we list the first report in the literature for each relevant algorithm.…”

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confidence: 99%

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

Ravenhill

Soday

Houghton

et al. 2020

Sci Rep

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“…This represents the first cell surface proteomic study in primary monocyte subsets. Up to now, other cell surface proteomic analyses in primary cells have been limited to CD4+ T-cells 16 , erythrocytes 17,18 , NK cells, adipocytes and a subset of tumour cells 19 . We quantify 373 cell surface proteins from classical monocytes, and further quantify 313 proteins across all monocyte subsets.…”

Section: Discussionmentioning

confidence: 99%

“…Data analysis was performed with modifications as previously described 18,38 . In the following description, we list the first report in the literature for each relevant algorithm.…”

Section: Discussionmentioning

confidence: 99%

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

Ravenhill

Soday

Houghton

et al. 2020

Preprint

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“…OMICs approaches(Nyalwidhe and Lingelbach, 2006;Ravenhill et al, 2019). The combination of our protocol with these techniques creates perspectives for more sophisticated ways of deciphering the ins and outs of host/pathogen interactions over the intra-erythrocytic cycle of Plasmodium on both the host and the parasite sides.Bio-protocol 10(11): e3647.…”

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Differential Fractionation of Erythrocytes Infected by Plasmodium berghei

Gnangnon¹,

Peucelle²,

Pierrot³

2020

BIO-PROTOCOL

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The study of host/pathogen interactions at the cellular level during Plasmodium intra-erythrocytic cycle requires differential extraction techniques aiming to analyze the different compartments of the infected cell. Various protocols have been proposed in the literature to study specific compartments and/or membranes in the infected erythrocyte. The task remains delicate despite the use of enzymes or detergents theoretically capable of degrading specific membranes inside the infected cell. The remit of this protocol is to propose a method to isolate the erythrocyte cytosol and ghosts from the other compartments of the infected cell via a percoll gradient. Also, the lysis of the erythrocyte membrane is done using equinatoxin II, which has proven to be more effective at erythrocyte lysis regardless of the cell infection status, compared to the commonly used streptolysin. The parasitophorous vacuole (PV) content is collected after saponin lysis, before recovering membrane and parasite cytosol proteins by Triton X-100 lysis. The lysates thus obtained are analyzed by Western blot to assess the accuracy of the various extraction steps. This protocol allows the separation of the host compartment from the parasite compartments (PV and parasite), leading to potential studies of host proteins as well as parasite proteins exported to the host cell.

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

Cited by 25 publications

References 61 publications

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

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

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

Differential Fractionation of Erythrocytes Infected by Plasmodium berghei

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