FINE MAPPING OF REGULATORY GENETIC VARIATION INVOLVING COMMON AUTOIMMUNE DISEASE ASSOCIATED HAPLOTYPES IN THE HUMAN MHC

Autoimmune diseases such as type I diabetes, multiple sclerosis and Crohn?s disease involve the body attacking itself through the immune system. Such diseases are common, particularly among younger people, for whom the consequences can be devastating. We know that autoimmune diseases occur more commonly in particular families and recent research has shown that many different genes are involved. In particular, scientists have found that a region of the human genome called the major histocompatibility complex on chromosome 6 is critically important in determining genetic susceptibility to autoimmune disease. We know that this part of our genome contains many genes involved in the immune response. The MHC is also a remarkably variable portion of the genome. It has proved very hard to define the particular genetic variants that are associated with developing disease within the MHC, or how such variants act to cause disease. In this research project we propose to investigate how genetic variation associated with disease affects the way genes are expressed. Genes are the blueprint of life, providing a code from which proteins are synthesized via an intermediary molecule called RNA. We will study how RNA is produced from DNA. We will do this in cells from the blood which are important in the immune response. We will study healthy volunteers who have different genetic backgrounds to see how this affects the process of gene expression. In particular we will study people with genetic backgrounds associated with autoimmune disease. We will also measure chemical changes in the DNA itself called methylation which can affect how genes are expressed. DNA methylation is an important control mechanism for the body to regulate expression of particular genes in particular cells at particular times. We know that this process may become dysregulated in disease and we plan to investigate this for genes in the MHC involved in autoimmunity. Overall our research will help scientists to better understand how genetic differences between people control the way our genes are regulated and define our risk of developing autoimmune diseases. This should help doctors to find new ways of treating these very important diseases and to target therapies to those who will benefit most. The work will be carried out at the University of Oxford. Our research will be published in scientific journals and promoted to the public through presentations at schools, the local and national media, and by providing information on the University website.

There is a significant inherited component in susceptibility to autoimmune disease with the most striking and robust genetic associations involving sequence variation in the major histocompatibility complex (MHC) on chromosome 6p21. However fine mapping these disease associations and resolving specific functional variants has proved a significant roadblock in the field. This is made more challenging at this locus due to the remarkably polymorphic nature of the MHC and extensive coinheritance or linkage disequilibrium between variants. There is also a need for a more integrated approach to the resolution of genetic determinants of common disease, recognising the importance of epigenetic mechanisms and context-specific regulation of gene expression. Here we propose to define functionally important genetic variants modulating gene expression for common autoimmune disease associated haplotypes spanning the MHC. We know that there is significant heritable variation in gene expression involving the MHC. Our data from lymphoblastoid cell lines HLA-homozygous for HLA-A1-B8-Cw7-DR3, A3-B7-Cw7-DR15 and A26-B18-Cw5-DR3-DQ2 demonstrates that haplotype-specific differences in gene expression are common and often involve differences in alternative splicing. We propose to extend this analysis to carry out transcript profiling using RNA-seq for B lymphocytes and monocytes from HLA-homozygous healthy volunteers selected by genotype for eight common autoimmune disease associated haplotypes. These haplotypes were completely re-sequenced as part of the MHC Haplotype Project and are strongly implicated in autoimmune diseases including type I diabetes, systemic lupus erythematosus and multiple sclerosis. We will then confirm haplotype-specific differences and define specific local (likely cis-acting) or distant (likely trans-acting) variants by expression quantitative trait mapping combined with allele-specific transcript quantification. Specific regulatory variants will be characterised in the context of the local regulatory landscape based on chromatin profiling together with reporter gene analysis, assays of protein-DNA interactions, small interfering RNAs and assays to define allele-specific expression of alternatively spliced isoforms. Our analysis of transcription will be complimented by epigenetic profiling to define haplotype-specific differences in DNA methylation involving the MHC. This work will provide unique insights into the functional basis of observed associations with autoimmune disease for commonly occurring haplotypes in European populations. This has significant translational importance in advancing our understanding of disease pathogenesis and defining patients at risk of disease or most significantly benefiting from specific therapies.