Characterization of the − 16C>G sequence variation in the promoters of both HBG1 and HBG2: Convergent evolution of the human γ-globin genes

We encountered a homozygous − 16C>G mutation in cis at identical positions in the promoters of both human γ-globin genes in a subject who was also homozygous for Hemoglobin C (HbC). Subsequent analysis of normal control individuals of African American ancestry revealed that both mutations were always present in cis with an allelic frequency of 3%. Furthermore, 10 out of 11 HbC subjects carried the − 16C>G sequence variations, suggesting an association with HbC. The − 16C>G mutation disrupts a putative CACCC box positioned between the TATA box and the transcriptional start site. However, the absence of high levels of HbF in HbC subjects homozygous and heterozygous for the − 16C>G sequence variation suggested no effect of this mutation on γ-globin gene expression in the adult stage of development. Further functional characterization by means of transient transfections in human erythroleukemic K562 cells showed that the − 16C>G promoter sequence variation did not have an effect on γ-globin expression in the fetal stage of development either. We therefore conclude that the − 16C>G γ-globin sequence variations are not beneficial to the clinical phenotype of HbC. The unique concurrent presence of this non-functional sequence variation is likely the result of a gene conversion event, and supports the concept of sequence homogenization between the two human γ-globin genes.