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5 promoter-driven reverse tetracycline-controlled transactivator mice (K5-rtTA) (Diamond et al., 2000). Bitransgenic mice (referred to as K5-Twist1 mice) showed speci?c expression of Twist1 protein in the basal epidermal layer upon doxycycline (dox) treatment (Figure S1C). Long-term induction of Twist1 alone in K5-Twist1 mice did not result in visible skin abnormalities (data not shown). To generate squamous cell carcinoma (SCC), K5-Twist1 mice and control single transgene littermates were treated with a single dose of 7,12-dimethylbenz[a]anthracene (DMBA) followed by weekly applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) for

20 weeks to allow skin tumor development (Abel et al., 2009;

Kemp, 2005;

Sun et al., 2007) (Figure 1A). At the end of TPA treatment, when all mice have developed multiple papil- lomas, we randomly divided these mice into two groups. One group of mice received doxycycline in the drinking water to allow continuous Twist1 expression in K5-positive tumor cells, even if tumor cells have migrated out of the skin and disseminated throughout the body. We used this systemic........