Figure 1.

HaCaT cells express ΔNp63α. (A) RT–PCR analysis of HaCaT cells exponentially growing in DMEM medium. Specific oligonucleotides for each of the six major isoforms were used with increasing PCR cycles. For details see Materials and Methods. (B) Western blot analysis of HaCaT nuclear extracts with the 4A4 antibody, which recognizes all p63 isoforms; a major band corresponding to the ΔNα isoform is predominant. (C) Immunofluorescence analysis of differentiation markers. HaCaT cells were grown in CaCl₂-free medium containing 10% foetal calf serum (FCS) (0 h, left panels) and in 1.2 mM CaCl₂, 0.1% FCS for three days (72 h, right panels). After fixation, cells were stained with anti-Cytokeratin 1, anti-Cytokeratin 8–18 differentiation markers, as well as anti-p63 antibodies. (D) RT–PCR analysis of p63 isoforms in HaCaT keratinocytes. mRNA was extracted from cells as in (C). Note that under these conditions, HaCaT cells tend to have higher levels of ΔNβ and γ isoforms. Nevertheless, the ΔNα isoform is still predominant.