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

Benjamin J Ravenhill,Jonathan M Goldberg,Robin Antrobus,Ambroise Ahouidi,Manoj T Duraisingh,Michael P Weekes,Tandakha Dieye,Usheer Kanjee,James Williamson,Luis Nobre

doi:10.1038/s42003-019-0596-y

Benjamin J Ravenhill, Jonathan M Goldberg + Show 8 more

Open Access

https://doi.org/10.1038/s42003-019-0596-y

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Abstract

Red blood cells (RBCs) play a critical role in oxygen transport, and are the focus of important diseases including malaria and the haemoglobinopathies. Proteins at the RBC surface can determine susceptibility to disease, however previous studies classifying the RBC proteome have not used specific strategies directed at enriching cell surface proteins. Furthermore, there has been no systematic analysis of variation in abundance of RBC surface proteins between genetically disparate human populations. These questions are important to inform not only basic RBC biology but additionally to identify novel candidate receptors for malarial parasites. Here, we use ‘plasma membrane profiling’ and tandem mass tag-based mass spectrometry to enrich and quantify primary RBC cell surface proteins from two sets of nine donors from the UK or Senegal. We define a RBC surface proteome and identify potential Plasmodium receptors based on either diminished protein abundance, or increased variation in RBCs from West African individuals.

Highlights

Red blood cells (RBCs) play a critical role in oxygen transport, and are the focus of important diseases including malaria and the haemoglobinopathies
We have previously shown that this approach has the power to quantify the majority of all cell surface proteins, identifying for example a critical drug transporter in cells latently infected with cytomegalovirus, in addition to a novel malaria receptor[6,27]
By using highly multiplexed tandem-mass tag (TMT)-based proteomics, the present study offered the opportunity to directly measure the variability in expression of whole RBC surface proteomes across multiple donors

Summary

Results

Optimised enrichment of RBC for proteomic analysis. Previous analyses of the RBC proteome have been complicated by the presence of contaminants in the RBC sample, including white blood cells, platelets and serum proteins. Previous studies have attempted to analyse the RBC membrane proteome by cellular lysis followed by isolation of the membrane fraction (‘white ghosts’)[25,28], or by metal ion chromoaffinity to deplete haemoglobin[16] Both are still subject to substantial contamination by abundant soluble intracellular proteins including haemoglobin, and lack specificity for surface proteins with extracellular domains. These include a number of proteins found at high abundance in human serum that are likely to be contaminants, for example complement component C1Q and apolipoproteins L1 and L2 (Supplementary Data 3b). 27 proteins were only identified in the present study (Supplementary Data 3e) Some of these are likely to represent previously undetected low-abundant RBC PM proteins, enriched by our plasma membrane-specific technique, including four multi-pass transmembrane solute carrier (SLC) transporters.

20 SLC40A1 10

Discussion

Methods