Ancient Evolutionary Origin and Positive Selection of the Retroviral Restriction Factor Fv1 in Muroid Rodents

Abstract

The laboratory mouse Fv1 gene encodes a retroviral restriction factor that mediates resistance to murine leukemia viruses (MLVs). Sequence similarity between Fv1 and the gag protein of the murine endogenous retrovirus L (MuERV-L) family of ERVs suggests that Fv1 was coopted from an ancient provirus. Previous evolutionary studies found Fv1 orthologs only in the genus Mus Here, we describe identification of orthologous Fv1 sequences in several species belonging to multiple families of rodents outside the genus Mus We show that these Fv1 orthologs are in the same region of conserved synteny, between the genes Miip and Mfn2, suggesting a minimum insertion time of 45 million years for the ancient progenitor of Fv1 Our analysis also revealed that Fv1 was not detectable or heavily mutated in some lineages in the superfamily Muroidea, while, in concert with previous findings in the genus Mus, we found strong evidence of positive selection of Fv1 in the African clade in the subfamily Muridae Residues identified as evolving under positive selection include those that have been previously found to be important for restriction of multiple retroviral lineages. Taken together, these findings suggest that the evolutionary origin of Fv1 substantially predates Mus evolution, that the rodent Fv1 has been shaped by lineage-specific differential selection pressures, and that Fv1 has long been evolving under positive selection in the rodent family Muridae, supporting a defensive role that significantly antedates exposure to MLVs.IMPORTANCE Retroviruses have adapted to living in concert with their hosts throughout vertebrate evolution. Over the years, the study of these relationships revealed the presence of host proteins called restriction factors that inhibit retroviral replication in host cells. The first of these restriction factors to be identified, encoded by the Fv1 gene found in mice, was thought to have originated in the genus Mus In this study, we utilized genome database searches and DNA sequencing to identify Fv1 copies in multiple rodent lineages. Our findings suggest a minimum time of insertion into the genome of rodents of 45 million years for the ancestral progenitor of Fv1 While Fv1 is not detectable in some lineages, we also identified full-length orthologs showing signatures of a molecular "arms race" in a family of rodent species indigenous to Africa. This finding suggests that Fv1 in these species has been coevolving with unidentified retroviruses for millions of years.

Keywords: Fv1 restriction factor; gammaretrovirus restriction; mouse leukemia viruses; rodent evolution.

FIG 1

An Fv1 ortholog is present in several rodent families. Shown is a cladogram illustrating the evolutionary relationship between rodent species with a genome assembly in the NCBI database. The species tree was generated with TimeTree (20). Genus, family, subfamily, and superfamily classifications for a subset of species are indicated on the right. + indicates the presence of an Fv1 ortholog or an Fv1 ORF as identified via BLAST searches of each genome assembly. The suggested positions of the Fv1 insertions into the rodent genomes are indicated by arrows. The solid arrow indicates the location of the minimum insertion time suggested by the database analysis, while the dashed arrow indicates the possible Fv1 insertion at the base of the suborder Myomorpha. *, the Fv1 ORF of Apodemus speciosus is split between two overlapping scaffolds. Merinones unguiculatus and Psammomys obesus contained only a partial ORF with premature stop codons.

FIG 2

Conserved synteny of Fv1. Genomic segments containing Fv1, Miip, and Mfn2 were extracted from the NCBI genome database for each indicated species. Cross-species homologies were analyzed using the MultiPipMaker alignment tool (25). Exons of Miip, Mfn2, and Fv1 are shown as black boxes in the mouse reference assembly. The Fv1 sequence is boxed in red. Regions with more than 50% but less than 75% identity are shown as green boxes. Regions with more than 75% identity are shown as red boxes. A cladogram representing the phylogenetic relationships between the species is shown on the left. The species tree was generated with TimeTree (20).

FIG 3

Phylogeny of Fv1 and Rbp3. Maximum-likelihood trees were generated with RaxML using aligned Fv1 (A) and Rbp3 (B) sequences from the indicated rodent species. Bootstrap values are shown next to each node. Each tree was rooted with Nannospalax galili. *, sample was originally identified as Mylomys dybowskii and then renamed Gerbilliscus sp. based on its Rbp3 sequence; **, sequence obtained from the NCBI database. Family and subfamily classifications for a subset of species are indicated on the right of each tree.

FIG 4

Evolution of Fv1 in African murids. A cladogram obtained using aligned Fv1 sequences from 13 African species of Muridae was generated with RaxML. The free-ratio model of PAML was used to calculate the dN/dS values shown above each branch with the replacement and synonymous changes in parentheses. The values on the branches under positive selection (dN/dS > 1) are highlighted in red. The asterisk indicates the only branch with a bootstrap value of <80. Genus, family, and subfamily classifications for a subset of species are indicated on the right of the tree.

FIG 5

Positive selection of Fv1 in African murids. A graphical representation of the Fv1 protein is shown, with highlighting to indicate the three variable regions previously identified as important in restriction (16). Amino acid residues that were identified as evolving under positive selection by four programs are shown above the Fv1 protein with differently colored arrows. Amino acid alignment of five segments of Fv1 in African murids (with Fv1 from the Mouse Reference Genome on top) are shown below, with residues under positive selection (as identified by PAML) shaded blue. The arrows below the alignments indicate previously identified positively selected sites in the genus Mus (17). The MHR of Fv1 is boxed.