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On the ancestry and evolution of the extinct dire wolf | bioRxiv

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On the ancestry and evolution of the extinct dire wolf

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https://www.biorxiv.org/content/10.1101/2025.04.09.647074v1

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bioRxiv

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Dire wolves ( Aenocyon dirus ) are extinct predators of Pleistocene North America. Although phenotypically similar to living wolves ( Canis lupus ), dire wolves have yet to be placed confidently in the canid family tree. We generated 3.4× and 12.8× paleogenomes from two well-preserved dire wolves dating to > 13,000 and > 72,000 years ago, and estimated consensus species trees for these and 10 canid species. Our results revealed that ∼2/3 of dire wolf ancestry is derived from a lineage sister to the clade comprising the gray wolf, coyote, and dhole, and the remaining ∼1/3 from a lineage near the base of Canini diversity. We identified 80 genes evolving under diversifying selection in dire wolves. Our results underscore the power of paleogenomes to resolve long-standing taxonomic questions and contribute to growing evidence of the role of post-speciation gene flow as an evolutionary force. ### Competing Interest Statement Authors affiliated with Colossal Biosciences and/or Form Bio may hold stock and/or stock options in these companies. E.K.K., C.E.M, B.vH., R.J.O., M.T.P.G, and L.D. are members of the scientific advisory board at Colossal Biosciences. M.S.S. is a consultant of Colossal Biosciences. G.R.R.M is an Investor and Cultural Advisor for Colossal Biosciences. G.C. is co-founder of Colossal Biosciences and others available at arep.med.harvard.edu/t The sequencing data underlying this study are available in the NCBI Sequence Read Archive, and can be accessed with BioProject accession PRJNA1222369. For the HyPhy analyses, we included a tarball of the multiple sequence alignment FASTAs of the coding sequences for all 80 genes under diversifying selection in the dire wolf branch as well as the HyPhy results JSON files from BUSTED, FitMG94, FEL, and MEME programs. These data and all supplemental tables are available in a Zenodo repository associated with this study under the BioRxiv DOI.

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On the ancestry and evolution of the extinct dire wolf

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Dire wolves ( Aenocyon dirus ) are extinct predators of Pleistocene North America. Although phenotypically similar to living wolves ( Canis lupus ), dire wolves have yet to be placed confidently in the canid family tree. We generated 3.4× and 12.8× paleogenomes from two well-preserved dire wolves dating to > 13,000 and > 72,000 years ago, and estimated consensus species trees for these and 10 canid species. Our results revealed that ∼2/3 of dire wolf ancestry is derived from a lineage sister to the clade comprising the gray wolf, coyote, and dhole, and the remaining ∼1/3 from a lineage near the base of Canini diversity. We identified 80 genes evolving under diversifying selection in dire wolves. Our results underscore the power of paleogenomes to resolve long-standing taxonomic questions and contribute to growing evidence of the role of post-speciation gene flow as an evolutionary force. ### Competing Interest Statement Authors affiliated with Colossal Biosciences and/or Form Bio may hold stock and/or stock options in these companies. E.K.K., C.E.M, B.vH., R.J.O., M.T.P.G, and L.D. are members of the scientific advisory board at Colossal Biosciences. M.S.S. is a consultant of Colossal Biosciences. G.R.R.M is an Investor and Cultural Advisor for Colossal Biosciences. G.C. is co-founder of Colossal Biosciences and others available at arep.med.harvard.edu/t The sequencing data underlying this study are available in the NCBI Sequence Read Archive, and can be accessed with BioProject accession PRJNA1222369. For the HyPhy analyses, we included a tarball of the multiple sequence alignment FASTAs of the coding sequences for all 80 genes under diversifying selection in the dire wolf branch as well as the HyPhy results JSON files from BUSTED, FitMG94, FEL, and MEME programs. These data and all supplemental tables are available in a Zenodo repository associated with this study under the BioRxiv DOI.

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