title: The AIRE -230Y Polymorphism Affects AIRE Transcriptional Activity: Potential Influence on AIRE Function in the Thymus.
abstract: BACKGROUND
  The autoimmune regulator (AIRE) is expressed in the thymus, particularly in thymic medullary epithelial cells (mTECs), and is required for the ectopic expression of a diverse range of peripheral tissue antigens by mTECs, facilitating their ability to perform negative selection of auto-reactive immature T-cells. The expression profile of peripheral tissue antigens is affected not only by AIRE deficiency but also with variation of AIRE activity in the thymus.
  
  
  RESULTS
  Therefore we screened 591bp upstream of the AIRE transcription start site including AIRE minimal promoter for single nucleotide polymorphism (SNPs) and identified two SNPs -655R (rs117557896) and -230Y (rs751032) respectively. To study the effect of these variations on AIRE promoter activity we generated a Flp-In host cell line which was stably transfected with a single copy of the reporter vector. Relative promoter activity was estimated by comparing the luciferase specific activity for lysates of the different reporter AIRE promoter-reporter gene constructs including AIRE-655G AIRE-230C, AIRE-655G AIRE-230T and AIRE-655A AIRE-230C. The analysis showed that the commonest haplotype AIRE-655G AIRE-230C has the highest luciferase specific activity (p<0.001). Whereas AIRE-655G AIRE-230T has a luciferase specific activity value that approaches null. Both AIRE promoter polymorphic sites have one allele that forms a CpG methylation site which we determined can be methylated in methylation assays using the M.SssI CpG methyltransferase.
  
  
  CONCLUSIONS
  AIRE-230Y is in a conserved region of the promoter and is adjacent to a predicted WT1 transcription factor binding site, suggesting that AIRE-230Y affects AIRE expression by influencing the binding of biochemical factors to this region. Our findings show that AIRE-655GAIRE-230T haplotype could dramatically alter AIRE transcription and so have an effect on the process of negative selection and affect susceptibility to autoimmune conditions.
authors.affiliation: School of Medicine and Biomedical Sciences, University of Sheffield, Sheffield, United Kingdom.
authors.forename: Thomas R J
authors.initials: TR
authors.lastname: Lovewell
authors.affiliation: Dermatology Department, Royal Hallamshire Hospital, Sheffield, United Kingdom.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: McDonagh
authors.affiliation: Dermatology Department, Royal Hallamshire Hospital, Sheffield, United Kingdom.
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Messenger
authors.affiliation: Department of Neuroscience, School of Medicine and Biomedical Sciences, University of Sheffield, Sheffield, United Kingdom.
authors.forename: Mimoun
authors.initials: M
authors.lastname: Azzouz
authors.affiliation: School of Medicine and Biomedical Sciences, University of Sheffield, Sheffield, United Kingdom.
authors.forename: Rachid
authors.initials: R
authors.lastname: Tazi-Ahnini
mesh_terms: D000483:Alleles; D001483:Base Sequence; D002460:Cell Line; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000235:genetics; D004847:Epithelial Cells / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics; D006239:Haplotypes / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D016415:Sequence Alignment; D013950:Thymus Gland / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics*; D024363:Transcription Initiation Site / Q000502:physiology; D014158:Transcription, Genetic / Q000235:genetics*; D000096927:AIRE Protein
chemical_list: D014157:Transcription Factors; D000096927:AIRE Protein