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Figure 4


Fig. 4. Keratin expression in mutant (Mut) and wild-type (WT) keratinocytes as judged by western blotting. The antibodies used are indicated on the left (K5, K14, K10, ß-actin). Total protein extracts were prepared from K1014chim and wild-type keratinocytes cultured in low calcium medium for 8 days. Expression of the differentiation marker K10 is not detectable in wild-type cells under these culture conditions. The signal obtained with the K10 antibody in K1014chim extracts therefore represents expression of the chimeric protein. Note that both the K10 and K14 antibody were raised against the C-termini of the keratins. Consequently, the K14 antibody does not detect its antigen in K1014chim extracts. ß-actin was used as a loading control to normalize keratin expression. K5 expression was similar in mutant and wild-type cells. As expected, the molar ratio of K5/K14 in wild-type cells was ~1:1. The ratio of K5/K1014chim in mutant cells was also calculated as ~1:1, indicating that the steady state levels of K14 (wild-type) and K1014chim (mutant) are similar.





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