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Sensing DNA? Aim for the cytoplasm in Systemic Lupus Erythematosus
Allyson Egan1, Jennifer Pacyna2, Margus Blagrove2, Michael Jones2 Department of Medicine1, Genomics Laboratory, MRC Clinical Sciences Centre2 Hammersmith Hospital, Imperial College London
Introduction
Aim2 is a novel cytoplasmic DNA sensor, which instigates the caspase pathway and formation of theinflammasome leading to cell death via IL1β and IL181. The gene encoding the protein is a member of the interferon response HIN200 family. Theproteins contain a HIN-200 amino acid region which binds to DNA and a pyrin domain which can homo/ oligodimerise2. Microarray data has reported over-expression of family member Ifi202 in the inbred Lupus prone mouse model BXSB and New Zealand subcongenic B6.Nba22. The telomeric region of chromosome 1 (C1) which contains the HIN200 locus in these two murine strains, along with a syntenic region in humans, has been linked with the development of Systemic Lupus Erythematosus (SLE) with phenotypic features of nephritis, autoantibody production and death2. A bioinformatic approach that includes comparative sequence analysis has been employed to investigate the homology and disparity in the genomic architecture at this locus.
Methods
C1 genomic DNA was isolated and fragmented from cells by the Wellcome Trust Sanger Institute. The random fragments were sequenced using Illumina Next Generation Sequencing technology to generate paired-end reads of 2x 75 bases. The fragments were aligned to the mouse reference genomeand viewed using the Integrated Genome Viewer (IGV version 2.2.5). Promoter, intronic and exonic sequences were scanned for single nucleotide polymporphisms (SNPs). Ensemble/ncbi databases, blast searches and clustral sequence comparisons were employed to investigate homology within the region. Real time PCR was performed to investigate differential expression ofAim2in autoimmune prone strain BXSB.
Results
The region is highly polymorphic. There are multipleSNPs identified in exonic, intronic and promoter regions in genes Ifi202, Ifi203, Ifi205, Mnda and Aim2. A lysine to glutamine switch occurs in Ifi202, Ifi203 and Ifi205. The start codon in Ifi203 contains a switch from methionine to threonine.Ifi205has altered charge due to glutamic acid replacement by glycine. In Mnda, glycine is switched to arginine along with a leucine switch to proline, which may cause alteration in protein structure. Exon 1 (E1) appears to be homologous in certain family members. A common E1 appears in full length in Mnda. Truncated versions occur in Ifi202 and Ifi204. The intact sequence demonstrates 98-100% homology with one another. Ifi205 demonstrates 81-83% homology with the other family members. AIM2 E1 shares only 17% homology with Mnda E1. In real-time PCR differential down regulation of expression has been identified in the autoimmune strains BXSB and subcongenicB10.Yaa.Bxs3 in comparison to the non-autoimmune control strain B10.Yaa.
Conclusion
A bioinformatic approach that includes comparative sequence analysis has identified homology between the gene families especially at E1,with the exception ofAim2. The lack of homology may reflect their different location and function within the cell. Furthermore, decreased expression of the cytosolic DNA sensor gene in the lupus prone mouse model BXSB and subcongenic B10.Yaa.Bxs3 in comparison to control B10.Yaahas been identified. Extensive polymorphic changes occurs in the region that result in amino acid substitutions which may alter protein structure.
References
1.An orthogonal proteomic-genomic screen identifies AIM2 as a cytosplasmic DNA sensor for the inflammasome. Burckstummer et al Nature Immunology vol 10, Number 3 March 2009. 2.Dissection of BXSB Lupus Phenotype using mice congenic for Chromosome 1 demonstrates that separate intervals direct different aspects of disease. Morley et al. JI 2004, 173:4277-4285