The tremendous phenotypic diversity of modern dog breeds represents the end point of a >15,000-year experiment in artificial and natural selection. Each breed has undergone strong artificial selection, in which dog fanciers selected for many traits including body size, fur type, color, skull shape, and even behavior, to create novel breeds. The adoption of the breed barrier rule that no dog may become a registered member of a breed unless both its dam and sire are registered members ensures a relatively closed genetic pool within each breed. As a result, there is strong phenotypic homogeneity within breeds including breed-associated genetic disease. This year we have made significant advances in our studies of canine bladder cancer, identifying a common druggable target in tumors. We have identified pathways that are mutated in canine transmissible tumors, which has allowed us to determine how tumors avoid immune surveillance. We have also continued our studies of canine origins. Canine Cancer Our recent genetic studies of dog disease have focused on cancer, which we argue is a strong model for human cancer genetic (Davis and Ostrander, 2015). With 20,000 new cases in the United States each year, canine invasive transitional cell carcinoma of the bladder (InvTCC) is a common, naturally occurring malignancy that shares significant histologic, biologic, and clinical phenotypes with human muscle invasive bladder cancer. In order to identify somatic drivers of canine InvTCC, we determined the complete transcriptome for multiple tumors by RNAseq (Decker et al., 2015). We found that all tumors harbored a somatic mutation that is homologous to the human BRAF(V600E) mutation, and an identical mutation was present in 87% of 62 additional canine InvTCC tumors. The mutation was also detectable in the urine sediments of all dogs tested with mutation-positive tumors. Functional experiments suggest that, like human tumors, canine activating BRAF mutations potently stimulate the MAPK pathway. Cell lines with the mutation have elevated levels of phosphorylated MEK, compared with a line with wild-type BRAF. This effect can be diminished through application of the BRAF(V600E) inhibitor vemurafenib. These findings set the stage for canine InvTCC as a powerful system to evaluate BRAF-targeted therapies, as well as therapies designed to overcome resistance, which could enhance treatment of both human and canine cancers. Canine Transmissible Venereal Tumor (CTVT). Canine transmissible venereal tumor (CTVT) is a parasitic cancer clone that has propagated for thousands of years via sexual transfer of malignant cells (Rev: Ostrander, Davis, Ostrander, Submitted). It is endemic everywhere except Antarctica. Because so little has been understood about the mechanisms that converted an ancient tumor into the world's oldest known continuously propagating somatic cell lineage, we created the largest existing catalog of canine genome-wide variation, which contains greater then 99% of canine genomic variation (Decker et al., 2015). We compared it against two CTVT genome sequences, thereby separating alleles derived from the founder's genome from somatic drivers of clonal transmissibility. We show that CTVT has undergone continuous adaptation to its transmissible allograft niche, with overlapping mutations at every step of immunosurveillance, particularly self-antigen presentation and apoptosis. We also identified chronologically early somatic mutations in oncogenesis- and immune-related genes that may represent key initiators of clonal transmissibility. Thus, we provide the first insights into the specific genomic aberrations that underlie CTVT's perseverance in canids around the world. A summary of our other cancer studies can be found in (Parker and Ostrander, 2014). Canine Genomics In the process of assembling 186 whole genome sequences from domestic dogs and wild canids, we realized that with little more work we could capture most of the variation present in modern dogs today. We now have a catalog of about 300 whole genome sequences that we are studying (Davis In Preparation). This is allowing us to identify genes and gene families important in canine domestication and breed development. Others share this interest and a number of collaborations have sprung up to attack this problem. We worked with (Freedman et al. 2014) To identify genetic changes underlying dog domestication and reconstruct their early evolutionary history. We generated high-quality genome sequences from three gray wolves, one from each of the three putative centers of dog domestication, two basal dog lineages (Basenji and Dingo) and a golden jackal as an outgroup. Analysis of these sequences supports a demographic model in which dogs and wolves diverged through a dynamic process involving population bottlenecks in both lineages and post-divergence gene flow. We narrow the plausible range for the date of initial dog domestication to an interval spanning 11-16 thousand years ago, predating the rise of agriculture. Regarding the geographic origin of dogs, none of the extant wolf lineages from putative domestication centers was found to be more closely related to dogs, and, instead, the sampled wolves form a sister monophyletic clade. This result, in combination with dog-wolf admixture during the process of domestication, suggests that a re-evaluation of past hypotheses regarding dog origins is necessary. Morphology A majority of our dog papers over the past 4-6 years reveal our growing understanding of canine genome organization and its relationship to morphologic variation between breeds. Our newest avenue of morphologic study is aimed at understanding the genetic underpinning of skull shape variation (Schoenebeck, 2014) which varies dramatically across breeds. To quantify the variation, we collected data from 533 museum skulls at 51 landmarks using a microscribe digitizer. The resulting principal components analysis (PCA) showed that the top four PCs account for about 77% of skull variance across breeds. We continue to make advances in this region and have written a review to summarize the state of the field (Schoenebeck and Ostrander 2014). Significant effort has also gone into developing a data repository of DNA and measurements and clinical information on Portuguese Water Dogs. This work is all being done in collaboration with K. Gordon Lark and Kevin Chase at the University of Utah. This remarkable breed is allowed considerable variation in body size and coat color. Over the years the Lark lab has built a tremendous resource of measurements and GWAS data that our own lab has continually benefitted from. We are now digging deeper into that dataset to begin studies of the genetics of metabolism and blood chemistries and to continue our work on morphology.
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