Interestingly, treatment of cultured keratinocytes with as little as 5 ng/ml of TGF-β2 caused a rapid and transient induction of Snail (Figure 5B). Following this treatment, Snail protein was detected within 30 min, peaked at 2 h, and then declined thereafter. The induction of Snail appeared to be specific for the active form of the growth factor, as pretreatment of TGF-β2 for 10 min at 100 °C obliterated the response [Figure 5B, lanes marked (–)]. By contrast, although TGF-β receptor activation remained elevated during the duration of the experiment (as measured by the sustained phosphorylation of the downstream effector SMAD2) Snail expression could not be maintained (Figure 5B). Thus, although Snail expression correlated with phosphorylated SMAD2 (pSMAD2) induction, its decline seemed to rely on secondary downstream events.