Mutations in Ampd3 were associated with a reduction in naive CD4⁺ T cells. (A) Percentage of naive CD4⁺ T cells determined by flow cytometry in 5 Ampd3 mutants created through ENU mutagenesis. Data points represent individual mice. Mean and standard deviation are indicated. The Manhattan plot shows −log₁₀(P value) (y-axis) plotted vs the chromosome positions of all mutations (x-axis) identified in the G₁ male founders containing the Ampd3 mutation. Horizontal pink and red lines represent thresholds of P = .05, without and with Bonferroni correction, respectively. VAR, REF, and HET mice descend from mutagenized mice and harbor additional ENU-induced mutations. (B) Genetic mutation, amino acid change, and the predicted Polymorphism Phenotyping v2 score (PPH) of all 5 Ampd3 mutant alleles. (C) Diagram of the protein domains of mouse AMPD3. The location of all 5 point mutations are shown. For comparison of expression, these mutant Ampd3 genes were tagged with a FLAG-tagged epitope and transfected into HEK293T cells. GFP serves as transfection control to demonstrate similar transfection efficiency among samples. AA, amino acid; HET, Ampd3^+/−; REF, Ampd3^+/+; WT, wild-type mice that do not descend from mutagenized mice; VAR, Ampd3^−/−.