FIG. 2.

Immunofluorescence analysis showing MK6a and MK6b expression patterns. (a through d) MK14 staining in green (FITC) and the overlapping MK6b staining in yellow (TxRed). (a) MK6b (yellow) is present in the companion cell layer (arrow) of telogen hair follicles in MK6a^−/− animals and MK6a^+/+ animals (inset). (b) MK6b (yellow) expression in the companion cell layer (arrow) of anagen hair follicles in MK6a^−/− and MK6a^+/+ mice (inset). Note that the outer ORS is only one cell layer thick in this section and only positive for MK14 (green); the outer ORS and the MK6b-positive companion cell layer are most readily distinguished in the lower part of the section (arrow). (c and d) MK6b is expressed in the periderm (yellow) of embryonic day 16.5 skin of MK6a^+/+ and MK6a^−/− animals. Note that the periderm is a single-cell layer covering the developing epidermis, which is highlighted by the MK14 antibody in green. (e and f) Samples taken from the edge of 34-h-old tape-stripping wounds. MK14 staining (TxRed) highlights the epidermis and the ORS of the hair follicles. (e) MK6 induction (yellow) in MK6a^+/+ animals is apparent in the entire epidermis and the ORS of the hair follicle. (f) MK6b in MK6a^−/− animals is induced in the suprabasal layers of the epidermis but not in the basal layer of the epidermis or the ORS (arrows). Note that MK6b staining is not seen in the companion cell layer of the follicle because the antibody used here for comparison with the MK6a^+/+ sample does not recognize MK6b in the companion cells of fixed tissue but recognizes it on unfixed frozen sections (not shown). (g) MK6a is expressed throughout the dorsal tongue epithelium in MK6a^+/+ animals (TxRed). The antibody recognizes both MK6a and MK6b. The overlapping yellow staining shows the suprabasal expression of MK16 in the papillae. Note that in the basal layers (arrow) MK6a (red) is expressed but not MK16. (h) MK6b (yellow) is restricted to the uppermost cells of the papillae in MK6a^−/− animals. The MK16 staining (green) in the MK6a^−/− tongue epithelium is identical to the MK16 staining seen in the MK6a^+/+ sample. (i) MK6b (yellow) is expressed suprabasally in the hard palate (counterstained with MK14; TxRed) in MK6a^−/− mice. (j) Suprabasal MK6b expression in MK6a^−/− footpad, which was counterstained with MK14 (TxRed). Note that MK6 expression in the footpad is patchy. (a, b, e, and f) Carnoys fixed samples; (c, d, and g to j) unfixed frozen sections. Scale bars, 50 μm.