Long-range haplotype analysis of the malaria parasite receptor gene ACKR1 in an East-African population

Qinan Yin,Willy Albert Flegel,Kshitij Srivastava,Addisalem Taye Makuria,Amha Gebremedhin

doi:10.1038/s41439-018-0024-8

Abstract

The human ACKR1 gene encodes a glycoprotein expressing the Duffy blood group antigens (Fy). The Duffy protein acts as a receptor for distinct pro-inflammatory cytokines and malaria parasites. We determined the haplotypes of the ACKR1 gene in a population inhabiting a malaria-endemic area. We collected blood samples from 60 healthy volunteers in Ethiopia’s southwestern low-altitude tropical region. An assay was devised to amplify the ACKR1 gene as a single amplicon and determine its genomic sequence. All haplotypes were resolved at 5178 nucleotides each, covering the coding sequence (CDS) of the ACKR1 gene and including the 5′- and 3′-untranslated regions (UTR), intron 1, and the 5′- and 3′-flanking regions. When necessary, allele-specific PCR with nucleotide sequencing or length polymorphism analysis was applied. Among the 120 chromosomes analyzed, 18 ACKR1 alleles were confirmed without ambiguity. We found 18 single-nucleotide polymorphisms (SNPs); only one SNP was novel. The non-coding sequences harbored 14 SNPs. No SNP, other than c.-67T>C, indicative of a non-functional allele, was detected. We described haplotypes of the ACKR1 gene in an autochthonous East-African population and found 18 distinct ACKR1 alleles. These long-range alleles are useful as templates to phase and analyze next-generation sequencing data, thus enhancing the reliability of clinical diagnostics.

Highlights

The human atypical chemokine receptor 1 gene (ACKR1, MIM #613665)[1,2] encodes a multi-pass transmembrane glycoprotein
A 12,125-nucleotide stretch of the ACKR1 gene was amplified as a single primary amplicon from 50 ng of genomic DNA using a long-range Taq polymerase (LongAmp Taq DNA Polymerase; New England Biolabs, Ipswich, MA, USA) and the first-round primers 5′-GCATTGCTTCCAGTTCTAAGCTC-3′ and 5′CGTCTCAATCGGTCCCTAAATCC-3′ (Eurofins MWG Operon; Huntsville, AL)
A random survey in 60 healthy volunteers was performed to describe the genetic variability of the ACKR1 gene for a large number of long-range haplotypes

Summary

Introduction

The human atypical chemokine receptor 1 gene (ACKR1, MIM #613665)[1,2] encodes a multi-pass transmembrane glycoprotein. It is a receptor for proinflammatory cytokines, such as interleukin-6 and -83,4, and the malaria parasites Plasmodium vivax and Plasmodium knowlesi[5,6,7]. The ACKR1 glycoprotein carries the five antigens of the Duffy blood group system (Fy)[12]. The two major antithetical antigens Fya and Fyb, encoded by the co-dominant alleles FY*A (FY*01) and FY*B (FY*02), are among the clinically most significant blood group antigens, involved in severe hemolytic transfusion reactions and hemolytic disease of the fetus and newborn[13,14,15,16,17]. FY*A and FY*B allele frequencies range from only 0 to 5% in East Africa[18,19]

Methods

Results

Conclusion