JCS003392 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. Alterations in the expression of adhesion proteins in Inv-Dsg2 mice. Indirect immunofluorescent staining of adult wild-type and transgenic skin using antibodies to desmosomal proteins (Dsg1-α, Dsg1-β, Dsg1-γ, Dsg3, desmoplakin and plakoglobin) and adherens junction proteins (E-cadherin and β-catenin). (A) Suprabasal expression of Dsg2 slightly enhanced the expression of desmosomal proteins but did not alter their expression pattern. Insets in i and k show wild-type desmoplakin (i) and plakoglobin (k) stainings obtained under the same exposure conditions used for transgenics (i-l) to further demonstrate the increase in their expression level in the Inv-Dsg2 transgenic skin. (B) Overexpression of Dsg2 did not dramatically alter the expression levels of E-cadherin or β-catenin. Nuclei were stained with DAPI (blue). DP, desmoplakin; PG, plakoglobin.

• Supplemental Figure 2 -

  Fig. S2. Effect of suprabasal Dsg2 on Triton-solubility and subcellular distribution of desmosomal proteins. Protein extraction was performed using adult wild-type and Inv-Dsg2 transgenic dorsal skin for Triton-soluble and −insoluble (A) and subcellular (soluble, high salt and insoluble; B) proteins. (A) Immunoblot analysis showed the presence of Dsg2-Flag in the Triton-soluble pool whereas other desmosomal proteins were found exclusively in the Triton-insoluble pool. Cytokeratin (CK) and α-tubulin levels were used as loading controls for the Triton X-100-insoluble and -soluble fractions, respectively. (B)Subcellular fractionation followed by immunoblotting showed Dsg2-Flag in all samples including the cytoplasmic (soluble) and nuclear (high salt) fractions. The asterisk (*) indicates a non-specific nuclear protein recognized by the Flag antibody. The nuclear pool of β-catenin was reduced in the Inv-Dsg2 transgenic compared to wild-type skin. α-Tubulin, γ-tubulin and cytokeratin served as loading controls for the cytoplasmic, nuclear and insoluble pools, respectively. S, Triton soluble; I, Triton insoluble.

• Supplemental Figure 3 -

  Fig. S3. Suprabasal expression of Dsg2 perturbs formation of the stratum corneum. (A) Hematoxylin and Eosin staining of dorsal skin of newborn wild-type and transgenic (Tg) littermates revealed disconnected and fragmented layers (arrows) in the stratum corneum (bracket) of Inv-Dsg2 mice. (B) Newborn wild-type and Inv-Dsg2 transgenic mice were tape stripped 10 times on the dorsal skin using CuDerm adhesive tapes. Tapes were adhered to microscope slides and photographed under phase contrast. TS-1, tape strip number 1; TS-10, tape strip number10.