Deep whole-genome sequencing based analysis of mosaic transposable mobile element insertions in adult human tissue. X. Zhu1, A. Fiston-Lavier2, D. Petrov3, M. Snyder4, D. Levinson1, A. Urban1,4 1) Psychiatry, Stanford University, Palo Alto, CA; 2) Computer Science, University of Montpellier2, Montpellier, Hérault, France; 3) Biology, Stanford University, Palo Alto, CA; 4) Genetics, Stanford University, Palo Alto, CA.

   Mobile elements (MEs) comprise a large portion of the human genome and some retain their capacity for transposition. Somatic retrotranspositions have been associated with several nervous system diseases, suggesting they may have a significant impact on the developing brain. To analyze somatic ME variation in human brain, previous studies have used target capture- or single cell-sequencing methods. Results suggest that somatic cell ME mutation is very likely, but it is not known how frequently new mobile element insertions (MEIs) occur and which proportions of cells, cell types or brain regions are affected. We developed an unbiased deep whole-genome sequencing-based approach to detect and quantify somatic MEIs without using target capture or whole-genome amplification methods for discovery. Using paired-end sequencing we sequenced DNA from postmortem brain from a subject with schizophrenia and a control subject, at mean 80x sequencing depth for superior temporal gyrus and 60x depth for both cerebellum and liver. We extensively modified an existing ME calling algorithm to identify novel MEIs. To validate predicted MEIs we used custom oligonucleotide capture for targeted re-sequencing of predicted novel MEIs at >300x depth. We identified 796 high-support potential novel MEIs each of which is present in only one of the four brain regions tested. The targeted-capture sequencing for validation tiled oligonucleotides across the regions surrounding ~20,000 of the whole-genome based MEI calls. Initial analysis showed high mappability of the resulting captured sequence and a substantial validation rate. At a false discovery rate below 10%, 82% of high-support Alu insertions and 73% of high-support LINE-1 insertions were validated. These validation rates will be further evaluated with digital droplet PCR, also to quantify the degree of mosaicism for each novel MEI. This project uses whole-genome sequencing to address more comprehensively the extent and precise genomic localization of somatic retrotransposition of mobile elements in human brain cells. This phenomenon represents an additional mechanism by which genomic variation can influence brain development and disease. Methods for analysis of novel ME sequences in deep whole-genome sequencing will be needed to fully characterize this type of mutation in diverse tissues and in subpopulations of cells using cell sorting techniques. This study is supported by grant R01MH094740 (to D.F.L.) from the NIMH.

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