PRESENTATION ABSTRACTS
breeds, plus small numbers of dogs from 7 additional breeds (42 diabetic cases, 92 controls). This led to further prioritization and genotyping of 2,500 variants in an additional cohort of Samoyeds and Labradors (57 cases, 73 controls). Notably, many diabetes-associated variants reside in genes involved in pancreatic beta-cell function or immune response and are distinct between breeds, confirming clinical observations that canine diabetes arises from multiple heterogeneous etiologies. Utilization of WGS to investigate complex traits such as canine diabetes will help to inform a precision medicine approach to these conditions. #23 Genomics and epigenomics of the feline Prader-Willi syndrome (PWS) orthologous-domain and generation of a genome-edited in vitro model Robert D. Nicholls 1 , Marie A. Johnson 1 , Leslie A. Lyons 2 , Susanne M. Gollin 3 , Erik A. Koppes 1 Robert.Nicholls@chp.edu 1 Division of Genetic and Genomic Medicine, Department of Pediatrics, UPMC Children’s Hospital of Pitts- burgh, and Univ. of Pittsburgh School of Medicine, Pittsburgh, PA, USA; 2 Department of Veterinary Med- icine & Surgery, College of Veterinary Medicine, Univ. of Missouri, Columbia, MO, USA; 3 Department of Human Genetics, Univ. of Pittsburgh Graduate School of Public Health, Pittsburgh, PA, USA Genomic imprints in eutherian mammals are epigenetic modifications highlighted by DNA meth- ylation that show parent-of-origin specific inheritance. However, no studies to date have evaluated genomic imprinting in Felis catus. After establishing that the human PWS-imprinted domain is highly conserved on F. catus chromosome B3, we demonstrated differential DNA methylation at the PWS-imprinting center (PWS-IC) accompanied by monoallelic expression of PWS-genes by studying the pseudodiploid Crandell-Rees feline kidney (CRFK) cell line. Next, the 2.2-kb PWS-IC was ablated by targeting Cas9 nuclease to CRISPR sgRNA sites flanking the PWS-IC. Many of 86 clonal lines recovered had genome-editing events (PWS-IC deletion, inversion, or scarring) on both alleles, including 16 with a deletion and 18 with an inversion. All eight lines having deletion or inversion of the unmethylated paternal PWS-IC lost expression of extended SNURF-SNRPN- snoRNA-lncRNA transcripts but retained expression of a proximal PWS three-gene cluster. In contrast, clonal lines with deletion or inversion of the methylated maternal PWS-IC allele had no effect on PWS gene expression. Intriguingly, several PWS-loci show features suggesting poly- morphism during Felidae evolution, while genomics characterization of SNURF-SNRPN-derived pseudogenes indicate features of germ cell-specific expression and function. In conclusion, study of PWS-IC genome-edited CRFK cell lines demonstrated strict monoallelic DNA methylation and that deletion or inversion of the unmethylated allele results in ablation of somatic cell expression of polycistronic PWS-gene expression downstream of the PWS-IC. Further study of genomic variation, evolution, and disease models of imprinted loci in carnivores are warranted.
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