The (A)gamma(+25 G -> A) polymorphism is associated with high HbF production leading to a decrease in binding activity of the LYAR transcription factor to the (A)gamma-globin gene in beta-thalassemia patients

Increase of gamma-globin genes expression and high level of fetal hemoglobin (HbF) in β-thalassemia patients is widely accepted as associated with a milder or even asymptomatic disease. The search for HbF-associated polymorphisms (such as the XmnI, BCL11A and MYB polymorphisms) has been object of several investigations for long time, but recently gained great attention, in order to stratify beta-thalassemia patients with respect to expectancy of the first transfusion, need for annual intake of blood, response to HbF inducers (the most studied of which is hydroxyurea). Agamma-globin gene sequencing was performed on genomic DNA isolated from a total of 75 beta-thalassemia patients, including 31 β039/β039, 33 β 039/β+IVSI-110, 9 β+IVSI-110/β+IVSI-110, one β0IVSI-1/β+IVSI-6 and one β039/β+IVSI-6. The results obtained show that a polymorphism (SNP) is present inbeta-thalassemia patients in the 5’UT sequence (+25) of the Agamma-globin gene. This was present in the NCBI-dbSNP (rs368698783), but it was not associated with β-thalassemia and any biological function. It the present study we have demonstrated that this polymorphism is linked with the Ggamma-globin-XmnI SNP (rs7482144). Moreover the SNP is associated with beta39-globin gene, but not with beta+IVSI-110-globin gene mutation. In addition, we confirmed by BIAcore analysis that this SNP alters the binding of LYAR (human homologue of mouse Ly-1 antibody reactive clone: a transcriptional repressor of the Agamma-globin gene) to the target 5’-GGTTAT-3’ site of the Agamma-globin gene. In agreement, we found that the Agamma(+25 G→A) and Ggamma-globin-XmnI SNPs are associated with high HbF levels in homozygous gamma039 thalassemia patients. Taken together, these data sustain the concept that the Ggamma-globin-XmnI(+)/Agamma-globin-(A) sequences are under genetic association with beta-thalassemia mutations