Localisation of cis regulatory elements at the beta-globin locus: analysis of hybrid haplotype chromosomes

Several cis elements at the beta-globin gene cluster and the upstream locus control region (LCR) have been implicated in modulation of fetal haemoglobin (Hb F) level in beta-globin disorders. To determine the role of elements at the LCR and the beta-globin gene cluster on HbF level among sickle cell anaemia (SCA) patients, hybrid haplotype betaS chromosomes exhibiting variation in the association of alleles of LCR hypersensitive site 2 (HS2) and the beta-globin gene cluster restriction fragment length polymorphosim (RFLP) haplotypes were identified in an unselected population of 100 patients. On 15 chromosomes the polymorphic HS2 short tandem repeat(TA)xN10-12(TA)y containing a Hox2 binding motif differed from that typically associated with the corresponding beta-globin gene cluster RFLP haplotype. Among patients homozygous for the Benin RFLP haplotype, in whom one chromosome carried the (TA)9N10(TA)10 allele, no effect on HbF level was observed. Polymorphism of the pre-Ggamma framework, an enhancer located 25 kb downstream of HS2 localised the breakpoint for each of these 'hybrid' haplotype chromosomes upstream of this element. Previously described hybrid haplotype chromosomes with the (TA)9N10(TA)10 HS2 allele associated with raised HbF by contrast arise by recombination 1 kb downstream of the pre-Ggamma framework. This study suggests that variability in HbF level associated with polymorphisn of the HS2 enhancer depend on downstream determinant (s) in tight linkage disequilibrium with HS2. The pre-Ggamma framework is the only known polymorphic cis-active determinant in this region.