Fig. 1.

FLG-luc2p mouse model. (a) Schematic diagram of the hFLG-10K-luc2p/Rosa26 knockout-replacement strategy used to generate the FLG-luc2p mouse model. The hFLG-10K-luc2p transgene contains a 10.6 kb human filaggrin promoter construct fused to the mammalian codon-optimized firefly luciferase gene luc2p. Single-copy C57BL/6J FLG-luc2p mice were generated via embryonic stem cell gene targeting into the murine Rosa26. (b & c) Luciferase expression patterns in FLG-luc2p^+/− mice were defined using in vivo bioluminescent imaging. Signals were strongest in the forepaws and hindpaws, although expression was detected in all skin samples monitored (see Supplementary Fig. 1). (d) WT (FLG-luc2p^-/-) and FLG-luc2p^+/− hindpaw tissues were hematoxylin/eosin (H&E) stained, or probed with α-keratin 1, α-filaggrin, or α-firefly luciferase antibodies and processed for immunofluorescence microscopy. FLG-luc2p reporter gene expression did not effect epidermal architecture (H&E) or alter endogenous K1 expression. Importantly, expression of luciferase and mouse filaggrin in the stratum granulosum confirmed that the hFLG-10K-luc2p transgene was appropriately expressed in the skin of FLG-luc2p^+/− mice. Scale bar = 50 µm. (e) In vivo imaging of FLG-luc2p^+/− mice (n = 12) at 24 hour intervals for 5 consecutive days revealed symmetric bioluminescent activity (%L/R ratio ≈ 101 ± 9) in the right and left paws at each time point. Color bar depicts luciferase light emission (LLE) intensity (photons/s/cm²/sr) all throughout. %L/R ratios were calculated throughout as follows: (left LLE/right LLE) × 100.