Abstract

Contrary to the mainstream blood group systems, P1PK continues to puzzle and generate controversies over its molecular background. The P1PK system comprises three glycosphingolipid antigens: Pk, P1 and NOR, all synthesised by a glycosyltransferase called Gb3/CD77 synthase. The Pk antigen is present in most individuals, whereas P1 frequency is lesser and varies regionally, thus underlying two common phenotypes: P1, if the P1 antigen is present, and P2, when P1 is absent. Null and NOR phenotypes are extremely rare. To date, several single nucleotide polymorphisms (SNPs) have been proposed to predict the P1/P2 status, but it has not been clear how important they are in general and in relation to each other, nor has it been clear how synthesis of NOR affects the P1 phenotype. Here, we quantitatively analysed the phenotypes and A4GALT transcription in relation to the previously proposed SNPs in a sample of 109 individuals, and addressed potential P1 antigen level confounders, most notably the red cell membrane cholesterol content. While all the SNPs were associated with the P1/P2 blood type and rs5751348 was the most reliable, we found large differences in P1 level within groups defined by their genotype and substantial intercohort overlaps, which shows that the P1PK blood group system still eludes full understanding.

Highlights

  • Despite great strides made to understand the molecular background of human blood groups, the P1PK blood group system continues to puzzle

  • The cells were analysed by flow cytometry and the antibody binding capacities (ABCs, the number of antibody molecules bound per cell) were calculated by interpolation from the calibration curve as described in the Single nucleotide polymorphisms toggle promiscuity of the Gb3/CD77 synthase and determine the P1PK blood type manufacturer’s protocol and based on the fluorophore-to-protein molar ratios of the FITCantibody conjugates

  • The manifold attempts to obtain a clear picture of how P1PK blood group differentiation ensues have been marred by the unorthodox activity of Gb3/CD77 synthase, the unusual ways in which genetic changes alter this enzyme’s activity, and the intertwined synthesis of the P1PK and other glycosphingolipid blood group antigens

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Summary

Introduction

Despite great strides made to understand the molecular background of human blood groups, the P1PK blood group system continues to puzzle. While it is well-established that the Pk antigen is synthesised by Gb3/CD77 synthase (α1,4-galactosyltransferase, P1/Pk synthase, encoded by A4GALT)[4], P1 has only recently been unequivocally shown to be a product of the same enzyme[5]. To address the controversy over allelic variations of A4GALT gene expression and P1/P2 phenotypic differentiation[14,15], we analysed the effect of four SNPs (rs8138197, rs2143918, rs2143919, rs5751348) previously reported to determine the P1/P2 status on A4GALT transcript levels and cellular-scale quantity of the P1 and NOR antigens in a sample of 109 NOR-negative and NOR-positive Polish individuals. We present the key results of this study in the form of univariate scatter plots, which faithfully represent distributions of continuous data, unlike still too often used bar plots[20]

Materials and methods
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