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

Benjamin J. Ravenhill,Robin Antrobus,Michael P. Weekes,Lior Soday,Jack Houghton

doi:10.1038/s41598-020-61356-w

Benjamin J. Ravenhill, Robin Antrobus + Show 3 more

Open Access

https://doi.org/10.1038/s41598-020-61356-w

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Journal: Scientific Reports	Publication Date: Mar 12, 2020
Citations: 27	License type: open-access

Affiliation: University of Cambridge

Abstract

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.

Highlights

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 were enriched by fluorescence activated cell sorting (FACS) after staining for CD86, CD14 and CD16 (Supplementary Fig. S1)
437 proteins were identified from the three samples, of which 373 were annotated ‘cell surface’, ‘plasma membrane’, ‘extracellular’ or with a short Gene Ontology term as previously described[13,14] (Supplementary Table S1d)

Summary

Introduction

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. Characterisation of cell surface proteins expressed by each subset is informative to improve understanding of phenotype, but may provide biological insights into function. We use highly multiplexed Tandem-Mass-Tag (TMT)-based mass spectrometry with selective cell surface biotinylation to characterise the classical monocyte surface proteome, interrogate the phenotypic differences between each monocyte subset to identify novel protein markers. 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[5,6,7,8]. Integrin alpha subunit 5 (ITGA5), complement receptor 1 (CR1/CD35) and Leukotriene B4 receptor (LTB4R) were defined as markers of classical monocytes, and Sialic Acid Binding Ig Like Lectin 10 (SIGLEC10) as a marker of non-classical cells

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