Identification of ATP2B4 Regulatory Element Containing Functional Genetic Variants Associated with Severe Malaria

Abstract

Genome-wide association studies for severe malaria (SM) have identified 30 genetic variants mostly located in non-coding regions. Here, we aimed to identify potential causal genetic variants located in these loci and demonstrate their functional activity. We systematically investigated the regulatory effect of the SNPs in linkage disequilibrium (LD) with the malaria-associated genetic variants. Annotating and prioritizing genetic variants led to the identification of a regulatory region containing five ATP2B4 SNPs in LD with rs10900585. We found significant associations between SM and rs10900585 and our candidate SNPs (rs11240734, rs1541252, rs1541253, rs1541254, and rs1541255) in a Senegalese population. Then, we demonstrated that both individual SNPs and the combination of SNPs had regulatory effects. Moreover, CRISPR/Cas9-mediated deletion of this region decreased ATP2B4 transcript and protein levels and increased Ca2+ intracellular concentration in the K562 cell line. Our data demonstrate that severe malaria-associated genetic variants alter the expression of ATP2B4 encoding a plasma membrane calcium-transporting ATPase 4 (PMCA4) expressed on red blood cells. Altering the activity of this regulatory element affects the risk of SM, likely through calcium concentration effect on parasitaemia.

ATP2B4 regulatory genetic variants are associated with mild malaria

“…Table 2 shows relevant Encode and GeneHancer regulatory elements [ 25 , 26 ], including those containing the studied SNPs. It should be stressed that these potential regulatory elements have been recently experimentally validated [ 17 , 19 ]. In particular, they were shown to have an enhancer activity, which increases the activity of the main ATP2B4 promoter [ 19 ].…”

“…It should be stressed that these potential regulatory elements have been recently experimentally validated [ 17 , 19 ]. In particular, they were shown to have an enhancer activity, which increases the activity of the main ATP2B4 promoter [ 19 ].…”

“…Mild malaria patient samples (n = 75) were taken at the Principal Hospital of Dakar and the regional hospital of Tambacounda in Senegal on the day of hospital admission [ 20 ], whereas control samples (n = 79) were taken from healthy individuals living in Dakar [ 19 ]. Additional file 1 : Table S1 shows the characteristics of malaria patients and control individuals.…”

“…The SNPs rs10751450, rs10751451, and rs10751452 that are in linkage disequilibrium with these tag SNPs were found to influence the expression of ATP2B4 [ 17 ], suggesting that they may be the causal SNPs. More recently, rs10900585, rs10751450, rs10751451, and rs10751452 were reported to be associated with severe malaria in a Senegalese population [ 19 ]. Furthermore, 5 new regulatory SNPs (rs11240734, rs1541252, rs1541253, rs1541254, and rs1541255) that disrupt the activity of a new regulatory element were also associated with severe malaria in the Senegalese population [ 19 ].…”

“…More recently, rs10900585, rs10751450, rs10751451, and rs10751452 were reported to be associated with severe malaria in a Senegalese population [ 19 ]. Furthermore, 5 new regulatory SNPs (rs11240734, rs1541252, rs1541253, rs1541254, and rs1541255) that disrupt the activity of a new regulatory element were also associated with severe malaria in the Senegalese population [ 19 ].…”