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Proc Natl Acad Sci U S A
Proc Natl Acad Sci U S A
PNAS
Proceedings of the National Academy of Sciences of the United States of America
0027-8424
1091-6490
National Academy of Sciences

202412585
10.1073/pnas.2412585121
letterLetterbiochemBiochemistry407
42
Letters
Biological Sciences
Biochemistry
An intronic polymorphism associated with 2,3-bisphosphoglycerate levels in human red cells is linked to expression of RhCE blood groups
McGowan Eunike C. a https://orcid.org/0000-0002-2098-4604

Storry Jill R. a b https://orcid.org/0000-0003-2940-2604

Olsson Martin L. martin_l.olsson@med.lu.se
a b 1 https://orcid.org/0000-0003-1647-9610

aDivision of Hematology and Transfusion Medicine, Department of Laboratory Medicine, Biomedical Center C14, Lund University, Lund SE-221 84, Sweden
bDepartment of Clinical Immunology and Transfusion Medicine, Office for Medical Services, Region Skåne, Lund SE-221 85, Sweden
1To whom correspondence may be addressed. Email: martin_l.olsson@med.lu.se.
22 8 2024
3 9 2024
22 8 2024
121 36 e2412585121Copyright © 2024 the Author(s). Published by PNAS.
2024
https://creativecommons.org/licenses/by-nc-nd/4.0/ This article is distributed under Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND).

Knut och Alice Wallenbergs Stiftelse (Knut and Alice Wallenberg Foundation) 501100004063 2020.0234 Martin L Olsson Vetenskapsrådet (VR) 501100004359 2019-02683 Jill R StorryMartin L Olsson Governmental grants to university healthcare in Region Skåne ALFSKANE-446521 Martin L Olsson access-typefree
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pmcWe read with interest how D’Alessandro et al. recently investigated the genetic underpinnings of the metabolic adaptations in red blood cells (RBCs) from healthy humans under high-altitude hypoxia (1). The authors concur previous in vivo studies in that 2,3-biphosphoglycerate (BPG) levels increase under high-altitude hypoxic conditions through their RBC metabolomics analysis of six healthy volunteers, who climbed to the highest city worldwide, La Rinconada, Peru (5,100 m above sea level). Strikingly, their RBC proteomics revealed the RhCE protein levels were most affected by ascent, acclimatization, and descent. Following this, metabolite quantitative trait loci analysis of 2,3-BPG using the omics and genotyping data from the Recipient Epidemiology and Donor Evaluation Study (REDS) was performed. A significant association between 2,3-BPG and genetic polymorphisms from chromosome 1 was found, and the polymorphism ranking highest in association was rs636889 located in intron 5 of RHCE.

The obvious question to ask is how a noncoding variant, rs636889, and/or other RHCE polymorphisms impact the RhCE protein to become a critical determinant of 2,3-BPG levels in RBCs of healthy volunteers? As D’Alessandro et al. mentioned, the RH genes comprise substantial genetic heterogeneity. We noted additional polymorphisms in linkage disequilibrium (LD) with rs636889 are present in the highly homologous RHD gene, which arose from a duplication event of RHCE (2). The RHD gene has single nucleotide variants (SNVs) distributed throughout the gene, including introns, depending on the phenotypic combination of Rh blood group antigens RhD, RhC, RhE, Rhc, and Rhe (3).

Similarly, in RHCE, more than one SNV can be used to predict the phenotype for C and c antigens, while one SNV in exon 5 defines the E and e antigens. LD analysis using LDlink (4) showed rs636889:C to correlate with SNVs encoding the RhC (rs586178:G, rs61777615:A) and Rhe (rs609320:C) antigens, whereas rs636889:T correlated with the Rhc (rs586178:C, rs61777615:G) and RhE antigens (rs609320:G; Fig. 1) (5, 6) in most populations. Lower linkage equilibria in African populations may be due to their unique diversity of RHCE alleles that affects the LD algorithm. Otherwise, in general, LDlink showed rs636889 travels with C/c and E/e antigen-encoding SNVs.

Fig. 1. Linkage disequilibrium patterns and allele frequencies from LDpop analysis for rs636889 and three SNVs implicated in expression of RhCE antigens. For each population, r2 values were plotted in a matrix heatmap using SRplot (7). The nucleotide changes linked with rs636889:C are underlined and those linked with rs636889:T are not. The nucleotide present in the reference allele for a given rs number is on the Left and the alternate allele on the Right.

Although all volunteers were reported as Rh-positive, their RhCE phenotypes were not specified (1). Interestingly, Rh antigen densities are known to vary depending on the combination of C, E, c, and e antigens (8). Computational modeling of these antigen combinations showed varying degrees of RhCE extracellular loop exposure, conformational structures, and interactions with the Rh-associated glycoprotein (RhAG) (9). With RhCE playing a structural role in the band 3-macrocomplex, a recognized participant of RBC oxygen regulation (10), it would be tempting to explore whether the RhD and RhCE phenotype plays a role in or can predict 2,3-BPG levels in the REDS study. Different RhCE protein isoforms could impact the structural stability or density of neighboring molecules in this macrocomplex implicated in ammonia or carbon dioxide transport.

In conclusion, we propose that an improved understanding of 2,3-BPG heterogeneity between humans may be gained by investigating the role of Rh phenotypes and their interactions within/around the macrocomplex.

We thank Ping Chun (Gloria) Wu from the Department of Laboratory Medicine and Sudip Ghosh from the Department of Experimental Medical Science, both at the Lund Stem Cell Center, Faculty of Medicine, Lund University, Lund, Sweden, for advice. This study was supported by the Knut and Alice Wallenberg Foundation (2020.0234 to M.L.O.), the Swedish Research Council (2019-01683 to M.L.O. and J.R.S.) and governmental ALF grants to the university healthcare in Region Skåne, Sweden (ALFSKANE-446521 to M.L.O.).

Author contributions

E.C.M. and M.L.O. designed research; E.C.M. performed research; E.C.M., J.R.S., and M.L.O. analyzed data; J.R.S. and M.L.O. supervised the study, edited the manuscript; and E.C.M. wrote the paper.

Competing interests

J.R.S. is the Senior Vice President for International Society of Blood Transfusion (ISBT) and on the ISBT Board of Directors. J.R.S. and M.L.O. has given educational lectures in exchange for honoraria from Biorad and QuidelOrtho. J.R.S. and M.L.O. are inventors on patents about blood group genotyping. J.R.S. and M.L.O. own 50% each of the shares in BLUsang AB, an incorporated consulting firm, which receives royalties for said patents.
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