Transmissible [corrected] dog cancer genome reveals the origin and history of an ancient cell lineage

Abstract

Canine transmissible venereal tumor (CTVT) is the oldest known somatic cell lineage. It is a transmissible cancer that propagates naturally in dogs. We sequenced the genomes of two CTVT tumors and found that CTVT has acquired 1.9 million somatic substitution mutations and bears evidence of exposure to ultraviolet light. CTVT is remarkably stable and lacks subclonal heterogeneity despite thousands of rearrangements, copy-number changes, and retrotransposon insertions. More than 10,000 genes carry nonsynonymous variants, and 646 genes have been lost. CTVT first arose in a dog with low genomic heterozygosity that may have lived about 11,000 years ago. The cancer spawned by this individual dispersed across continents about 500 years ago. Our results provide a genetic identikit of an ancient dog and demonstrate the robustness of mammalian somatic cells to survive for millennia despite a massive mutation burden.

Figure 1. CTVT tumors, karyotypes and copy number

(A) Samples sequenced in this study. Both tumor (24T, 79T) and host (24H, 79H) DNA was sequenced from the two individuals shown. (B) Multiplex FISH using red fox probes to investigate karyotypes of a normal female dog (left) and CTVTs collected in Cape Verde (center) and Italy (right). (C) CTVT genomic copy number for 24T (upper panel) and 79T (lower panel). Red and blue points represent total copy number and minor copy number (i.e. copy number of the allele present in fewer copies) respectively calculated using normalized read counts at each of 2,544,508 SNP loci. Chromosomes are represented by horizontal alternating black and gray bars.

Figure 2. CTVT mutations

Analyses were performed on a set of 395,306 CTVT variants that were annotated as somatic due to their heterozygous status within genomic regions that have undergone both loss of heterozygosity (LOH) and duplication. (A) Simple mutation spectrum in CTVT. Mutations are labelled in pyrimidine context. (B) Dinucleotide mutation spectrum in CTVT. The “first base” was defined as the mutation with the lower chromosome coordinate and the “second base” is immediately adjacent to the first base on the same strand. The strand is displayed relative to the pyrimidine context of the first base. 3,518 dinucleotide mutations were included in the analysis. (C) The proportion of mutations in CTVT explained by mutational signatures A to D.

Figure 3. Tracing the CTVT Founder Animal

(A) Principal component analysis of 1,106 wolves, dogs and coyotes using genotypes at 23,782 polymorphic SNP loci (8, 9). Each individual is represented by a single colored dot and positions of CTVT (inferred from the genotypes of 24T and 79T), 24H and 79H are indicated. Breeds were classified as Modern or Ancient according to (27). (B) Positions of CTVT (left), 24H (center) and 79H (right) on pairwise distance tree comparing genotypes at 23,782 SNP loci with 1,106 other dogs, wolves and coyotes (8, 9). Only the closest breeds to CTVT, 24H and 79H are shown. Breeds containing members that most strongly clustered with CTVT and 79H after genotype resampling are marked with red text and * (see Table S5, 24H did not cluster strongly with individuals from any single breed). NGSD, New Guinea Singing Dog. (C) Sex chromosome copy number of 24H (a female dog), 79H (a male dog), 24T and 79T determined by counting the number of reads aligning to X and Y chromosomes and normalized to 79H. Y chromosome reads in 79T are likely to be derived from contaminating host DNA. (D) Proportion of annotated SNP loci in germline diploid regions that are heterozygous in 24H, 79H, 24T and 79T. (E) Timeline for CTVT origin and divergence.