Fig. 5

MuERV-L(ancML) replication can be inhibited by innate immune effectors. a Enumeration of G418 resistant colonies generated in the presence of increasing amounts of mouse IFNα. CHO cells were transfected with plasmids expressing a neo gene (NEO), L1.3, or ancML and cultured with increasing amounts of mouse IFNα for 2 days before G418 selection. Data are mean ± SD from 3 independent experiments. b Infectivity of MuLV on CHO cells expressing Fv1^bbn. Percentage of MuLV infected (GFP positive) cells in CHO cells stably expressing Fv1^bbn (red) or an empty vector (black). Circles and triangles indicate infection by N-tropic or B-tropic MuLV, respectively. c Enumeration of G418 resistant colonies of CHO cells expressing Fv1^bbn or an empty vector were transfected with plasmids expressing a neo gene (NEO), L1.3, or ancML. Data are mean ± SD from 2 independent experiments. d Representative images of Immunofluorescence assays on CHO cells stably expressing an HA-tagged version of mouse APOBEC3 or an empty vector. CHO cells were fixed with 4% PFA and stained with anti-HA antibodies. e Enumeration of G418 resistant colonies of CHO cells expressing mouse APOBEC3. Three clones of CHO cells expressing HA-tagged mA3 or an empty vector were transfected with plasmids expressing a neo gene (NEO), L1.3, or ancML. Data are mean ± SD from 3 experiments with independent single cell clones. f and g Analysis of MuERV-L elements using Hypermut 2.0. Ratio of G to A mutations at preferred mA3 editing sites (RD 3′ to a G) (Y-axis) plotted against ratio of G to A mutations at disfavored mA3 editing sites (YN|RC 3′ to a G) (control ratio, X-axis). No 5′ context was imposed (f), or sites with a 5′ C to the mutated G were excluded (g). 230 gag–pol containing MuERV-L elements in the mouse genome were compared to their consensus sequence. Data points in red and orange indicate MuERV-L sequences that were statistically significantly enriched in putative mA3 induced mutations (p value < 0.05). Data points in orange represent MuERV-L elements that are statistically significantly enriched mA3-induced mutations in both analyses (p value < 0.01). h Profile of G to A transitions in two putatively mA3-edited MuERV-L proviral sequences compared to a consensus sequence. The profile of the reference MuERV-L sequence is shown for comparison (ML_ref, non significantly mA3 edited). Lines in red and cyan represent putative mA3-derived G to A transitions, not accounting for the + 2 position (dinucleotide changes from GG to AG and GA to AA respectively), whereas lines in green and magenta represent non mA3-derived G to A transitions (GC to AC and GT to AT respectively). Lines in yellow indicate gaps compared to the consensus sequence