Ancient origin of the furin sequence in the wolf F8 gene

Background: Our previous characterization of canine coagulation factor VIII (FVIII) in the naturally occurring hemophilia A (HA) dog models provided insight into the evolution of the FVIII protein. We noted a unique sequence (H1645) in the R-X-X-R furin-recognition motif within the B domain of canine FVIII that was distinct from the sequence (R1645) in humans and other model organism species, including mice, rats, pigs, and sheep. R1645 was associated with lower secretion and biological activity, which can be improved with the canine H1645 sequence. Aim: Herein, we sought to determine the evolutionary origin of the canine H1645. Methods: Gray wolves (Canis lupus) are the ancestors of the domestic dogs (C. lupus familiaris). Thus, we searched for H1645 in members of the genus Canis, including the gray wolf and the closely related coyote (C. latrans), in several free-range animals from diverse geographic locations and compared it to several breeds of domestic dogs. We also compared our sequences to publicly available reference sequences for other members of the class Mammalia, order Carnivora, and suborder Caniformia. Results: 19 X chromosomes from 12 gray wolves, at least 20 X chromosomes from 14 coyotes, and at least 12 X chromosomes from 12 domestic dogs of at least 10 distinct breeds all had the canine H1645 variant. Other members of the order Carnivora and suborder Caniformia, whose sequences are publicly available, had the R1645 sequence. Conclusion: Our results suggest that the H1645 variant in the furin-consensus sequence was likely derived after the infraorders Cynoidea and Arctoidea diverged but before the separation of the gray wolf and coyote and persists through the domestic dog.