Blood disorders such as β-thalassemia and sickle-cell anaemia arise due to mutations in the genes encoding the β-globin subunits of haemoglobin. Haemoglobin is a tetrameric heterodimer composed of two α-like and β-like subunits present in red blood cells that is essential for carrying oxygen around the body. To cater for varying oxygen demands during development, the composition of haemoglobin changes in a tightly-regulated process known as globin switching. This involves two switches in the composition of β-globin, first from embryonic-to-foetal globin during gestation, and secondly from foetal-to-adult after birth. Due to compelling evidence suggesting that increased foetal-globin can compensate for loss of functional adult-globin, studies have aimed to uncover the mechanisms behind globin switching as a means to ‘reawaken’ the foetal genes therapeutically.
GWAS studies have identified three major regions that are associated with high foetal-globin levels, however, these sites cannot account for the variation in foetal-globin expression between humans. In order to expand our knowledge on the variants involved with foetal-globin regulation, we aim to identify additional genetic variants that are involved with the regulation of globin genes. Regions of interest include those within the β-globin locus, such as the 5’HS4-LCR polymorphism, as well as in the genes encoding factors known to be important in globin regulation, such as Klf1. Our approach is to engineer variants of interest into erythroid progenitor cell lines by CRISPR/Cas9 in order to characterise the impact of these variants on foetal-globin expression. Future work will involve analysis of globin expression at the mRNA level by RT-PCR and at the protein level by flow cytometry.
We aim to utilise our growing understanding of the importance of these genetic variants to aid in informing novel therapeutic approaches to the treatment of blood disorders such as β‑thalassemia and sickle-cell anaemia.