Red cell and iron metabolism: using genetic and functional studies and CRISPR/Cas9 to identify of causal variants for blood cell phenotypes
One of our major interests is the role of genetic traits that determine resilience or susceptibility to the sequelae of blood donation. We are therefore particularly interested in traits that maintain iron homeostasis and also the biology of side effects that follow iron deficiency including neurocognitive syndromes.
The aim of this project is to understand the role of individual genetic variation in determining blood cell traits and iron metabolism, and more specifically, how these traits and mechanisms define the ability to donate, and the quality of red cells and other haematopoietic cells. In this project, you will use the existing and novel genetic data to define how genetic traits determine variation in red blood cell phenotype and/or iron metabolism.
As a starting point, we will leverage the genetic association data from a recent genome-wide association study (GWAS) of 30 different red blood cell, platelet and leukocyte indices in nearly 200,000 participants from the UK Biobank and INTERVAL cohorts. Our comprehensive analysis has identified two thousand genetic loci robustly associated with major blood cell phenotype(s) (Astle et al, 2016). In addition, we have recently completed a major RCT of 50,000 blood donors, randomised to different donation intervals (de Angelantonio et al, 2017). We have used the results of this trial, biomarkers of iron metabolism and extended genotypic analyses from all participants to complete a GWAS of restless legs syndrome (Schormair et al, 2017) and are completing new GWAS of iron metabolism including the effects of genetic traits on the ability to give blood frequently and levels of ferritin and hepcidin (Raymond et al., in preparation).
The challenge now is to define the causal variants and the functional impact of these novel loci, as well as to pinpoint the relevant genes and regulatory networks. Many novel missense mutations have been associated with red blood traits In order to interpret the functional mechanisms driving the observed genetic associations, we will first examine the function implications of several novel missense mutations that have been associated with red blood traits. For example, one intriguing set of associations with multiple hematopoietic lineages was of variants in genes involved in sphingosine signaling. In addition, our preliminary analyses also indicate novel loci that modulate iron uptake and link iron metabolism with other cellular phenotypes.
Other recent work indicates that many associated variants lie in non-coding sequence, suggesting that some of the causal variants must alter how genes are switched on and off. We will therefore integrate cell type-specific epigenome and gene expression data and results from chromatin conformation capture in red blood cell precursors, accessory cells and hepatocytes to examine the role of non-coding variants.
In these studies, we will use stem cells and erythroblast lines, macrophage and hepatocyte lines and determine phenotype, transcriptome and the proteome of respective cells before and after knockdown of the candidate gene using siRNA and CRISPR/Cas9. We will also use CRISPR/Cas9 gene editing to introduce the putative functional genetic variant. Finally, we will exploit the recall-by-genotype capabilities of the INTERVAL bioresource and use a wide variety of functional assays in primary cells to confirm and extend in vitro observations.
Students will have access to a wide-range of seminars and training opportunities through the many research institutes and centres based in Oxford. Students are also able to attend the Methods and Techniques course run by the MRC Weatherall Institute of Molecular Medicine. This course runs through the year, ensuring that students have the opportunity to build a broad-based understanding of differing research techniques.
Generic skills training is offered through the Medical Sciences Division's Skills Training Programme. This programme offers a comprehensive range of courses covering many important areas of researcher development: knowledge and intellectual abilities, personal effectiveness, research governance and organisation, and engagement, influence and impact. Students are actively encouraged to take advantage of the training opportunities available to them.
The Department has a successful mentoring scheme, open to graduate students, which provides an additional possible channel for personal and professional development outside the regular supervisory framework. We hold an Athena SWAN Silver Award in recognition of our efforts to support the careers of female students and staff.
Astle et al., (2016) The allelic landscape of human blood cell trait variation and links to common complex disease, Cell, 167, 1415–1429.
Di Angelantonio E, Thompson SG, Kaptoge S, Moore C, Walker M, Armitage J, Ouwehand WH, Roberts DJ*, Danesh J*; INTERVAL Trial Group. (2017) Efficiency and safety of varying the frequency of whole blood donation (INTERVAL): a randomised trial of 45 000 donors. Lancet, Sep 20. pii: S0140-6736(17)31928-1.
Schormair S et al., (2017) Meta-analysis of > 100,000 individuals of European ancestry identifies 19 genome-wide significant risk loci for restless legs syndrome . Lancet Neurol, 16, 898-907.