Thesis

Gene expression profilling in a family with a novel form of beta-thalassemia

The genetic causes of β-thalassemia are largely well described. However, the disease is very heterogeneous at both the molecular and clinical levels. Studying the transcriptome profiles of β-thalassemia patients, especially individuals who carry novel mutations in the β-globin gene (HBB), may improve our understanding of the heterogeneity and molecular mechanisms of the disease and its possible treatment. Here, I characterized members of a family with β-thalassemia using whole genome expression analysis. I report a novel mutation in the exon 1 region of HBB (HBB:c.51C>T) that was associated with an unexpected phenotype of β-thalassemia in a heterozygote who also carries a typical β-thalassemia allele. I analyzed effects of the novel mutation at the transcriptome level by RT-qPCR and high-throughput RNA sequencing using an Illumina Hiseq2500 system. The results revealed that the novel mutation creates a cryptic donor splice site in the HBB, which causes alternative splicing from the site and down-regulates (~0.7) expression of the β-globin. Gene expression profiling analysis showed that there were more than 300 differentially expressed genes (DEGs) in β-thalassemic blood. The DEGs were enriched in pathways that are directly or indirectly related to β-thalassemia such as hemopoiesis, heme biosynthesis, response to oxidative stress, inflammatory responses, immune responses, controlling of Circadian rhythms, apoptosis, and other cellular activities. It was possible to compare these findings with published results of RNA-seq analysis of sickle cell disease and KLF1-null anemia, and recognize similarities and differences in their transcriptional expression patterns. While many DEGs involved in response to hemolysis, iron homeostasis, and anemia were in common between these three types of anemias, over 200 DEGs were unique to β-thalassemia. Although this study was limited by the small sample size of the patients, it provides a wealth of data on β-thalassemia because it is the first broad investigation of blood cell gene expression in this disease, and gives us novel insight that can be used in drug discovery to identify novel therapeutic approaches for the disease.

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