Extracting Cell-type-specific Gene Expression Differences from Complex Tissues

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OR.19. CD48 Deficiency Precipitates Autoimmune Glomerulonephritis in Lupus Prone Mice Elahna Paul1, Yvette Latchman 2, Andrew Kirby1, Anna Koh1, Sarah Njoroge1, Marianela Feliu1, Arlene Sharpe 3, Mark Daly1, Robert Colvin1. 1Massachusetts General Hospital, Boston, MA; 2University of Washington School of Medicine, Seattle, WA; 3Brigham and Women's Hospital, Boston, MA Sle1b is a lupus susceptibility locus on mouse chromosome 1 that is associated with autoantibody production. This region contains members of the SLAM/CD2 gene family including CD48. In the context of a permissive Sle1b haplotype, CD48 deficiency promotes fatal glomerulonephritis (GN). This observation prompts the hypothesis that CD48 protects lupus prone individuals from progressive autoimmune disease. CD48-deficient C57BL/6 mice (B6. CD48-/-) spontaneously develop proliferative lupus GN. Early disease is characterized by mesangial expansion and immune complex deposition, followed by mesangial proliferation, leukocyte infiltration and crescent formation. Progressive fibrosis culminates in renal failure. B6.CD48-/mice also have autoantibodies. In contrast, CD48-/-BALB/c mice and CD48-/-F1 [B6 × BALB] progeny are free of both autoantibodies and renal disease. One backcross of F1 animals to the B6.CD48-/- parental strain restores autoantibody production (without nephritis) in enough N2 progeny to implicate a mendelian modifier of this humoral trait. Renal disease, however, is recovered only after additional backcrosses to the B6.CD48-/- strain, demonstrating that GN pathogenesis is polygenic. Genome wide SNP and QTL analyses of N2 animals have linked autoantibody production to the MHC region of chromosome 17. Mapping of the nephritic phenotype is underway. In summary, CD48-deficient mice spontaneously develop lupus-like disease and fatal GN. Presumably in combination with 129 alleles of the Sle1b locus on chromosome 1, permissive genetic modifiers of disease lie within the C57BL/6 (versus BALB/c) genome and include sites linked to the murine MHC. These data suggest that CD48 is a crucial participant in multigenic interactions that can precipitate lethal autoimmune disease. doi:10.1016/j.clim.2010.03.036

OR.20. Extracting Cell-type-specific Gene Expression Differences from Complex Tissues Shai Shen-Orr, Alexander Gaidarski, Robert Tibshirani, Purvesh Khatri, Mark Davis, Atul Butte. Stanford University, Stanford, CA Blood is a complex tissue made up of many different celltypes, each with its own functional attributes and molecular signature. The proportions of any given cell-type in the blood can vary markedly, even between normal individuals. This results in a significant loss of sensitivity in gene expression studies of blood cells and great difficulty in identifying the cellular source of any perturbations. Ideally, one would like to perform differential expression analysis between patient groups for each of the cell-types within a tissue but this is

Abstracts impractical and prohibitively expensive. We describe here a statistical methodology which enables the analysis of gene expression in complex tissues, such as blood, at a much higher resolution than previously possible. Given microarray data from biological samples, it estimates the relative cellfrequency of each cell-type in the sample, deconvolves the gene expression measurements for each cell-type, and uses this to identify differentially expressed genes between groups at a cell-type specific level. We apply our method to a large collection of blood gene expression measurements from a variety of diseases. This allows the identification of a large number of differentially expressed genes in specific cell-types that are otherwise undetectable. We characterize immune cell-state for each disease and classify diseases by their gene expression similarity at the cell-type specific level. doi:10.1016/j.clim.2010.03.037

OR.21. Deep Resequencing of SLE Susceptibility Loci in a Population of SLE Patients Edward Wakeland1, Ekta Rai1, Benjamin Wakeland 1, Chaoying Liang1, Nancy Olsen1, David Karp1, Jennifer Kelly 2, Swapan Nath 2, John Harley 2, Patrick Gaffney 2. 1 University of Texas Southwestern Medical Center, Dallas, TX; 2 University of Oklahoma, Oklahoma City, OK To identify the functional variations within the N 20 susceptibility loci that contribute to SLE, we are using targeted genomic resequencing to characterize SLE-associated LD blocks in 600 SLE genomes. Initial studies utilized custom oligonucleotide slide arrays (Roche NimbleGen 380K) to capture non-repetitive genomic sequences from 52 SLE-associated genomic segments. Sequence data for 32 Caucasian SLE patients were obtained using genome sequence capture protocols and sequencing methodology on the Illumina GAIIx Genome Analyzer. Of the N40 gigabases of total sequence reads aligning to the human genome in this study, 60-80% mapped to the targeted regions, yielding 35-45-fold average coverage. A comparison of 50 SNP genotypes previously determined in 5 samples with these sequences revealed concordance N 0.95. Our ongoing analyses have identified extensive SNP variations and CNVs in SLE susceptibility alleles. For instance, in BANK1, ITGAM, LY9 and BLK, we have detected from 35-83% (100-536 SNPs) of the total variations reported for these genes in the dbSNP 129 database. Out of these variations, 515% are located in regions that potentially impact gene function, consistent with the presence of extensive functional variations in SLE susceptibility alleles. This ongoing genomic sequencing analysis is being combined with RNASEQ analysis of EBV cell lines derived from these patients to identify causative genetic variations for SLE. We anticipate that N 80 SLE susceptibility genomes will be resequenced in the next two months. These data will be presented and discussed. doi:10.1016/j.clim.2010.03.038