Transcriptome Sequencing of Human Aging Brain Tissue Uncovers Widespread Genetic Effects on Splicing Alternations in Alzheimers disease. T. Raj1,2,3,4, J. Xu1, C. McCabe3, J. A. Schneider5, N. Pochet1,3,4, D. A. Bennett5, P. L. De Jager1,2,3,4 1) Department of Neurology, Brigham and Womens Hospital, Boston, MA; 2) Department of Medicine, Division of Genetics, Brigham and Womens Hospital, Boston, MA; 3) The Broad Institute, Cambridge, MA; 4) Harvard Medical School, Boston, MA; 5) RUSH Alzheimers Disease Center, RUSH University Medical Center, Chicago, IL.

   Recent studies suggest that alternations in RNA processing might be a key mechanism in Alzheimers disease (AD) pathogenesis. Here, we used RNA-sequencing to survey genome-wide transcriptome profiles from dorsolateral prefrontal cortex in 541 subjects of a longitudinal age-related cognitive decline cohort. To explore the mechanisms that drive transcriptional variation associated with cognitive decline, we performed a systematic QTL study aimed at identifying local and distal genetic effects on exons, genes, and isoforms in the brain transcriptome. We identified hundreds of RNA splicing events that correlate with individuals trajectory of age-related cognitive decline. We observe widespread genetic effects on gene and isoform expression levels, with variants influencing transcript ratios (tr-QTLs) of 2,651 (FDR 0.10) genes in cis and 264 (bonferroni p< 0.05) genes in trans. Of these, ~15 % of tr-QTLs are retained-intron transcripts subject to non-sense mediated decay, suggesting an overabundance of unspliced transcripts in the aging brain that is consistent with recent reports. We also detect several AD susceptibility alleles influencing isoform ratios in cis including genes involved in immune processes (PILRB, C4A, and C4B) and others (PICALM and ABCA7). These results include a variant in the sortilin-related VPS10 protein (SORCS3) associated with isoform ratios of over 17 genes in trans, some of which are located within AD-associated loci including SORL1, CLU, and PILRA. Interestingly, we identified six trans-QTL effects on isoform ratios mediated through a cis-association with a known splicing factor hnRNP C: the SNP is associated with decreased expression of hnRNP in cis and splicing of the six target genes. At a less stringent threshold, we observe similar pattern of altered regulation of RNA targets mediated by other splicing factors including CUG-BP1 (CELF1) and SR proteins that are targets of cis-eQTL. We plan to pursue UV Cross-Linking and Immunoprecipitation (CLIP) experiments in the same tissue to validate the downstream RNA targets of the splicing factors. Overall, our comprehensive study of the aging brain transcriptome provides evidence that (1) genetic variants, including AD susceptibility variants, regulate the generation of mRNA isoforms through effects on RNA splicing factors and (2) the splicing machinery is altered by the pathophysiology of AD.

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