Paleovirology: Using Endogenous Retroviruses Within Animal Genomes To Understand The Deep History Of Retroviruses

Retroviruses infect a wide range of vertebrates. The understanding of the deep history and host distribution of retroviruses remains far from complete. Retroviruses can be integrated into their host genomes and occasionally become vertically inherited genomic loci. These integrated retroviruses, known as endogenous retroviruses (ERVs), provide "molecular fossils" for past retroviral infections and are useful for studying the deep history and ecology of retroviruses. ERVs are highly abundant in vertebrate genomes. However, endogenous foamy viruses and lentiviruses appear to be extremely rare. The primary focus of the research presented here is to discover and analyze novel endogenous foamy viruses and lentiviruses in animal genomes. Foamy virus has been thought to exclusively infect three placental mammal superorders (Laurasiatheria, Euarchontoglires, and Xenarthra). The discovery of endogenous foamy viral elements (CoeEFV) in the genome of the coelacanth (Latimeria chalumnae) extends the host range of foamy viruses to fish lineages (Appendix A). I demonstrate that foamy viruses have likely codiverged with their vertebrate hosts for more than 407 million years. The discovery of CoeEFV provides evidence for an ancient marine origin of retroviruses. Endogenous foamy virus-like elements (PSFVaye) were also identified within the genome of a Malagasy lemur, the aye-aye (Daubentonia madagascariensis) (Appendix B). Phylogenetic analysis shows that PSFVaye is divergent from all currently known simian foamy viruses, suggesting a potentially ancient association between foamy viruses and primate species. Another novel endogenous foamy virus (CaEFV) was identified in the genome of the Cape golden mole (Chrysochloris asiatica). The discovery of CaEFV reveals foamy virus infection in the placental mammal superorder Afrotheria and the long-term cospeciation between foamy viruses and placental mammals (Appendix C). Lentivirus has been thought to have a relatively recent origin. Endogenous lentivirus insertions (MELV) were discovered within the genomes of some species of the Weasel family (Mustelidae) (Appendix D). I verified the presence of MELV insertions in the genomes of several species of the Lutrinae and Mustelinae subfamilies but not the Martinae subfamily, which suggests that the lentiviral invasion likely occurred between 8.8 and 11.8 million years ago. Phylogenetic analysis suggests MELV might represent a novel lentiviral group. The discovery of MELV extends the host range of lentiviruses to the Caniformia. Endogenous lentiviruses (GvaELV) were also identified in the genome of the Sunda flying lemur (Galeopterus variegatus) (Appendix E). Phylogenetic analysis shows that GvaELV is a sister group of all known lentiviruses. The discovery of GvaELV might give a clue to the early evolution of lentiviral genome architecture. In summary, the discoveries and analyses of these novel ERVs provide important insights into the deep history and ecology of foamy viruses and lentiviruses as well as the retroviruses as a whole

The Aspergillus fumigatus Damage Resistance Protein Family Coordinately Regulates Ergosterol Biosynthesis and Azole Susceptibility

Ergosterol is a major and specific component of the fungal plasma membrane, and thus, the cytochrome P450 enzymes (Erg proteins) that catalyze ergosterol synthesis have been selected as valuable targets of azole antifungals. However, the opportunistic pathogen Aspergillus fumigatus has developed worldwide resistance to azoles largely through mutations in the cytochrome P450 enzyme Cyp51 (Erg11). In this study, we demonstrate that a cytochrome b5-like heme-binding damage resistance protein (Dap) family, comprised of DapA, DapB, and DapC, coordinately regulates the functionality of cytochrome P450 enzymes Erg5 and Erg11 and oppositely affects susceptibility to azoles. The expression of all three genes is induced in an azole concentration-dependent way, and the decreased susceptibility to azoles requires DapA stabilization of cytochrome P450 protein activity. In contrast, overexpression of DapB and DapC causes dysfunction of Erg5 and Erg11, resulting in abnormal accumulation of sterol intermediates and further accentuating the sensitivity of ΔdapA strains to azoles. The results of exogenous-hemin rescue and heme-binding-site mutagenesis experiments demonstrate that the heme binding of DapA contributes the decreased azole susceptibility, while DapB and -C are capable of reducing the activities of Erg5 and Erg11 through depletion of heme. In vivo data demonstrate that inactivated DapA combined with activated DapB yields an A. fumigatus mutant that is easily treatable with azoles in an immunocompromised mouse model of invasive pulmonary aspergillosis. Compared to the single Dap proteins found in Saccharomyces cerevisiae and Schizosaccharomyces pombe, we suggest that this complex Dap family regulatory system emerged during the evolution of fungi as an adaptive means to regulate ergosterol synthesis in response to environmental stimuli