A remarkable case of HbH disease illustrates the relative contributions of the α-globin enhancers to gene expression

Understanding fundamental principles of enhancer biology at a model locus

“…One patient homozygous for a deletion of R2 had a moderately severe anaemia showing that the remaining enhancers can drive sufficient α‐globin gene expression to sustain a relatively normal life, supporting the idea that R1 together with R3 and R4 can act as ‘shadow’ enhancers, seemingly redundant enhancers but shown to add robustness to tissue‐ and developmental‐specific gene expression. [ 94,111 ] Despite careful analysis of ∼50 individuals with the phenotype of α‐thalassaemia and yet no associated deletions or insertions, we have never observed a deleterious single nucleotide polymorphism in the R2 enhancer. Although such polymorphisms may exist, it is likely that they do not cause sufficient change in α‐globin expression to cause a recognisable change in phenotype.…”

Functional analysis of three new alpha-thalassemia deletions involving MCS-R2 reveals the presence of an additional enhancer element in the 5’ boundary region

“…The MCS-R1 element has been supposed to contribute 10% of α-globin gene expression. Its sequence analysis ruled out the presence of mutations that could impact on the transcriptional rate of the (αα)CT and (αα)FG deletions (S2 Fig) . To acquire useful data on the function of specific sequences, we compared the phenotype of the three new deletions with that of three short deletions newly described, removing almost only the MCS-R2 element and part of the upstream boundary region: (αα)#18 (6710 bp starting from -3500); (αα)ALT (3361 bp starting from -300); the (αα)JX (742 bp starting from -333) [10,[16][17][18][19][20]. The extent of the deletions and the heterozygote's phenotype, (Fig 1 and Table 2) support the hypothesis of an enhancer activity present in the boundary region upstream of MCS-R2.…”

“…All these data allow us to conclude that both (αα)JX and (αα)ALT deletion are able to support the synthesis of α-globin genes in a higher way than (αα)FG; in other words, we can say that (αα)FG delimits further the region showing an enhancer activity, in accordance with a more severe phenotype. As shown in Fig 1B the (αα)FG spans 350 bp, compared to the shorter deletion (αα)JX, the upstream sequence required for full α-globin genes expression [10,[16][17][18][19][20].…”

“…Quantitative chromosome conformation capture (q3C) highlighted a looping model in which multiprotein complexes at the MCS-R2 and the gene promoters collide, and their interactions become stabilized, in erythroid cells [ 9 ]. The relative contributions of the α-globin enhancers to globin gene expression in the human has been suggested to be ~ 10% (MCS-R1), 90% (MCS-R2), 2% to 3% (MCS-R3), and 2% to 3% (MCS-R4) [ 5 , 10 ].…”