Fig. 4. Keratinocyte differentiation is accompanied by transcription upregulation of the sumoylation system. HaCaT cells maintained in low [Ca²⁺] medium (Time 0) were induced to differentiate by replacing the medium with high [Ca²⁺] medium. RNA was extracted at various times post Ca²⁺ induction as indicated. The extracted RNA was analyzed for expression of differentiation marker genes (A,B) or sumoylation system genes (C-E), and the mRNAs were detected either by quantitative RT-PCR (A,C,E) or RT-PCR (B,D). For the quantitative RT-PCR, the 0 hour time samples were set to a value of 1, and the values at other time points are relative to the 0 hour value. For the RT-PCR, 18S rRNA was used as the internal standard. The quantitative results in A,C and E were the average of at least three independent experiments.