Allele-selective RUNX1 binding regulates P1 blood group status by transcriptional control of A4GALT.

Westman JS, Stenfelt L, Vidovic K, Möller M, Hellberg Å, Kjellström S, Olsson ML

Blood 131 (14) 1611-1616 [2018-04-05; online 2018-02-08]

P1 and P k are glycosphingolipid antigens synthesized by the A4GALT-encoded α1,4-galactosyltransferase, using paragloboside and lactosylceramide as acceptor substrates, respectively. In addition to the compatibility aspects of these histo-blood group molecules, both constitute receptors for multiple microbes and toxins. Presence or absence of P1 antigen on erythrocytes determines the common P1 (P1+Pk+) and P2 (P1-Pk+weak) phenotypes. A4GALT transcript levels are higher in P1 individuals and single-nucleotide polymorphisms (SNPs) in noncoding regions of A4GALT, particularly rs5751348, correlate with P1/P2 status. Despite these recent findings, the molecular mechanism underlying these phenotypes remains elusive. The In(Lu) phenotype is caused by Krüppel-like factor 1 (KLF1) haploinsufficiency and shows decreased P1 levels on erythrocytes. We therefore hypothesized KLF1 regulates A4GALT expression. Intriguingly, P -specific sequences including rs5751348 revealed potential binding sites for several hematopoietic transcription factors, including KLF1. However, KLF1 binding did not explain 1 P -specific shifts in electrophoretic mobility-shift assays and small interfering RNA silencing of 1KLF1 did not affect A4GALT transcript levels. Instead, protein pull-down experiments using P but not 1 P oligonucleotide probes identified runt-related transcription factor 1 (RUNX1) by mass spectrometry. Furthermore, RUNX1 binds 2 P alleles selectively, and knockdown of 1 RUNX1 significantly decreased A4GALT transcription. These data indicate that RUNX1 regulates A4GALT and thereby the expression of clinically important glycosphingolipids implicated in blood group incompatibility and host-pathogen interactions.

Targeted and Structural Proteomics [Service]

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PubMed 29438961

DOI 10.1182/blood-2017-08-803080

Crossref 10.1182/blood-2017-08-803080