Fig. 1: Phenotype of mice encoding an APLAID mutation in Plcg2 (p.Ser707Tyr).
a, Scheme of Flp-excision at the Plcg2 recombined locus. Breeding was established with C57BL/6 Flp deleter mice to excise the Neomycin selection cassette and to generate heterozygous mice carrying the Neo-excised point knock-in mutation p.Ser707Tyr. b, Plcg2S707Y/+ mice displayed cutaneous lesions on paws, ears and tail. c, Severity of skin inflammation is reflected by an APLAID skin score of 2 on a 0–5 scale after weaning up to 24 d of age (Plcg2+/+, n = 19; Plcg2S707Y/+, n = 5; at 2–3 weeks of age). d, A growth curve exhibits stunted weight gain of Plcg2S707Y/+ mice (Plcg2+/+, n = 14; Plcg2S707Y/+, n = 9; at 2–4 weeks of age). e, Plcg2S707Y/+ mice (n = 5) showed splenomegaly compared to their Plcg2+/+ littermate controls (n = 7) at 6 weeks of age. f, Kaplan–Meier analysis demonstrated survival rates of 6 weeks in Plcg2S707Y/+ mice after weaning (Plcg2+/+, n = 5; Plcg2S707Y/+, n = 6; at 4–6 weeks of age). g, At 6 weeks of age, histopathological examination demonstrated myeloid immune cell infiltration in paws, ears, tails, lung, gut and spleen. One representative immunohistochemistry (IHC) section (MPO+ and F4/80+) from three independent experiments is shown. h, During disease onset, data obtained from ADVIA blood cell analyzer revealed an increase of neutrophils and a mild decrease in lymphocytes (Plcg2+/+, n = 7; Plcg2S707Y/+, n = 4; at 2 weeks of age). i,j, FACS analysis showed an increase of myeloid cell counts in the skin and lung at disease onset, while neutrophil numbers were reduced in the BM (n = 3 mice per genotype; at 2 weeks of age). k, During disease, peak neutrophil counts normalized in the blood as assessed by ADVIA analyzer (Plcg2+/+, n = 5; Plcg2S707Y/+, n = 6; at 6 weeks of age). l,m, FACS analysis of the skin and lung demonstrated increased numbers of myeloid cell numbers as APLAID progressed, while myeloid numbers in the BM remained unaltered (n = 3 mice per genotype; at 6 weeks of age). n,o, FACS analysis of lymphoid cells displayed normal T cells and reduced B cells (n = 3 mice per genotype; at 5 weeks of age). p, Quantification of immunoglobulin subtypes measured by ELISA revealed increased IgG2a levels of Plcg2S707Y/+ mice (n = 3; at 6 weeks of age) in comparison to Plcg2+/+ littermate controls (n = 8; at 6 weeks of age). Error bars represent the mean ± s.e.m. Statistical significance for skin score was determined by a two-way analysis of variance (ANOVA) with Bonferroni post-test correction. Statistical significance for the survival curve was determined by a Mantel–Cox test. Spleen weights, longitudinal weight data and cell numbers between two groups were determined by a two-sided unpaired Student’s t-test.
