Abstract
Hemolysis as a suggested cause of hyperuricemia is based upon the fact that red blood cells contain uric acid. The development of hyperuricemia also generates from an increased synthesis of nucleic acids occurring as part of the erythropoietic response to hemolysis in hemoglobin disorders such as sickle cell anemia, α-thalassemia, and β-thalassemia. In addition, multiple genome-wide association studies (GWAS) have reported significant association between uric acid levels and specific genomic loci. However, the mechanism of hyperuricemia still remains controversial and it is also unknown whether African Americans have higher prevalence of hyperuricemia due to genetic vs environmental risk factors. Here, we used joint admixture mapping and association testing to identify genetic variants associated with serum uric acid levels in African American. Interestingly, we detected 6 SNPs (rs2855125, rs2855126, rs11036415, rs11036496, rs4320977, and rs4348933) in an intergenic region of the β–globin cluster on chromosome 11 are associated with high levels of serum uric acid in populations of African ancestry. Next, we explored the potential regulatory role of intergenic SNPs associated with hyperuricemia using luciferase reporter gene activity assays and electrophoretic mobility shift assays (EMSA). Each SNP-containing DNA fragment was amplified by PCR using human genomic DNA and inserted into a firefly luciferase reporter vector, pGL3-basic vector. 293T or K562 cells were co-transfected with these constructs and a Renilla luciferase vector to control for transfection efficiency. Expression of firefly luciferase driven by each SNP-containing DNA fragment was measured by a dual luciferase reporter assay and normalized by Renilla luciferase expression. SNPs rs2855126, rs11036496 and rs4348933 on chromosome 11 had significantly greater expression levels of firefly luciferase than pGL3-basic-transfected cells in both 293T and K562 cells. Of these, the SNP rs2855126 ancestral C allele (associated with higher serum uric acid levels) showed significantly higher luciferase activity than the derived G allele. Functionally, the luciferase activities from these constructs were determined to be very similar in both cell lines used. Alleles altering expression were further assessed for binding of nuclear extracted proteins by EMSA. We found specific gel shift bands for SNPs rs2855126, rs11036496, and rs4348933, suggesting these SNPs are situated in the binding site of potential transcription factors. These data provide new insights into the potential contribution of imbalanced β-globin gene expression to hyperuricemia. DisclosuresNo relevant conflicts of interest to declare.
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