Genetic variation of the whole ICAM4 gene in Caucasians and African Americans

Abstract

Background: Landsteiner-Wiener (LW) is the human blood group system Number 16, which comprises two antithetical antigens, LW(a) and LW(b) and the high-prevalence antigen LW(ab) . LW is encoded by the intracellular adhesion molecule 4 (ICAM4) gene. The ICAM4 protein is part of the Rhesus complex in the red cell membrane and is involved in cell-cell adhesion.

Study design and methods: We developed a method to sequence the whole 1.9-kb ICAM4 gene from genomic DNA in one amplicon. We determined the nucleotide sequence of Exons 1 to 3, the two introns, and 402-bp 5'-untranslated region (UTR) and 347-bp 3'-UTR in 97 Caucasian and 91 African American individuals.

Results: Seven variant ICAM4 alleles were found, distinct from the wild-type ICAM4 allele (GenBank KF712272), known as LW*05 and encoding LW(a) . An effect of the LW(a) /LW(b) amino acid substitution on the protein structure was predicted by two of the three computational modeling programs used.

Conclusions: We describe a practical approach for sequencing and determining the ICAM4 alleles using genomic DNA. LW*05 is the ancestral allele, which had also been observed in a Neanderthal sample. All seven variant alleles are immediate derivatives of the prevalent LW*05 and caused by one single-nucleotide polymorphism (SNP) in each allele. Our data were consistent with the NHLBI GO Exome Sequencing Project (ESP) and the dbSNP databases, as all SNPs had been observed previously. Our study has the advantage over the other databases in that it adds haplotype (allele) information for the ICAM4 gene, clinically relevant in the field of transfusion medicine.

Figure 1. ICAM4 protein and ICAM4 pre-mRNA

Schematic models of ICAM4 protein are depicted for the 3 known isoforms (upper panel). Variations were found at 3 amino acid positions (red circles). The first 22 amino acid positions are predicted to be a signal peptide (blue circles). Additional predicted structural features are the 2 immunoglobulin (Ig) domains (yellow and orange circles) and a transmembrane segment (brown circles). Isoform Long (1) is a single-pass transmembrane protein (A), while Isoform Short (2) and Isoform 3 are secreted (B and C). Some protein segments in isoforms 2 and 3 differ from isoform 1 (purple, pink and green circles). The projections on circle surfaces denote the positions of the 4 N-glycosylation sites. The 7 variant nucleotide positions are depicted in the 3 pre-mRNA ICAM4 isoforms (bottom panel). Three variants were found in the exons (boxes) and 4 in the introns (lines). The nucleotide stretches of the exons are colored according to the encoded protein segments. The exon boundaries in the ICAM4 cDNA, as reflected in the amino acid sequence, are indicated (black bars).

Figure 2. Phylogeny of ICAM4 alleles in humans

A phylogenetic tree of ICAM4 is shown for the 8 alleles found in this study using the ICAM4 sequence from Pan troglodytes (NC_006486.3) for external rooting. Clustering of the described ICAM4 alleles is based on Neighbor-Joining method. For each evolutionary step, the event is indicated; depicted distances of the alleles are arbitrary.