, 2-way ANOVA with Bonferroni's test for multiple comparisons (G and K), or Mann-Whitney test (L). *P < 0.05; **P < 0.01; ***P < 0.001. Scale bars: 50 μm; original magnification in B and I, ×20; NS, not significant. The Journal of Clinical Investigation R E S E A R C H A R T I C L E4 4 9 9 jci.org Volume 126 Number 12 December 2016 some expression of keratin 14 (K14) in epithelial cells. However, these organs did not exhibit histological abnormalities (Supplemental Figure 3). These results indicate that the cause of death in such mice is more likely related to the skin abnormalities. Because of the accelerated mortality in Rabgef1 K-KO mice, and to assess the function of keratinocyte-intrinsic RABGEF1 in adult mice that would exhibit no potential developmental effects of constitutive Rabgef1 deletion, we generated mice in which Rabgef1 can be depleted in keratinocytes by tamoxifen treatment, i.e., K14-Cre-ERT2 + Rabgef1 fl/fl mice (called Rabgef1 KERT2-KO mice). Topical tamoxifen treatment on the back skin of adult Rabgef1 KERT2-KO mice, but not adult Rabgef1 fl/fl mice, promoted the development of skin lesions that were similar to those in Rabgef1 K-KO mice and were associated with RABGEF1 deletion in keratinocytes (Figure 1, H-K). Such skin lesions were associated with a modest, albeit statistically significant, increase in the number of bacteria that could be cultured from the affected skin ( Figure 1L), but not with evidence of associated systemic bacterial infection (Supplemental Figure 4).RABGEF1 deletion in keratinocytes results in epidermal barrier dysfunction, features of type 2 skin inflammation, and elevated levels of IgE antibodies. Histological and immunohistological analyses of back skin from newborn mice did not reveal obvious differences between Rabgef1 K-KO and control mice (Supplemental Figure 5). By contrast, back skin lesions of postnatal day 7-9 as well as day 49-56 (i.e., adult) Rabgef1 K-KO mice exhibited marked epidermal hyperplasia (Figure 1, F and G) and spongiosis ( Figure 2A These observations prompted us to investigate whether keratinocyte-intrinsic RABGEF1 deficiency resulted in epidermal barrier dysfunction. We used our model of inducible keratinocyte Rabgef1 deletion by topical tamoxifen treatment of Rabgef1 KERT2-KO mice and assessed barrier function by measuring transepidermal water loss (TEWL) (ref. 36 and Figure 2F). As shown in Figure 2G, TEWL was significantly increased in the back skin of tamoxifen-treated Rab gef1 KERT2-KO mice versus tamoxifen-treated control mice. Consistent with the increased permeability of the skin, the transepidermal passage of a harmless antigen, ovalbumin (OVA), patched on the skin was also potentiated when RABGEF1 was defective in keratinocytes. Indeed, dermal DC uptake and endosomal digestion of OVA was also substantially induced in tamoxifen-treated Rabgef1 KERT2-KO mice, whereas OVA was less accessible to DCs in vehicle-treated Rab gef1 KERT2-KO mice or tamoxifen-treated littermate control mice ( Figure 2H).(DCs) with the capacity to induce a ...

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