TGF-β and its signaling mediators, Smad2, -3, and -4, are involved with tumor suppression and promotion functions. Smad4 -/-mouse epidermis develops spontaneous skin squamous cell carcinomas (SCCs), and Smad3 -/-mice are resistant to carcinogen-induced skin cancer; however, the role of Smad2 in skin carcinogenesis has not been explored. In the present study, we found that Smad2 and Smad4, but not Smad3, were frequently lost in human SCCs. Mice with keratinocyte-specific Smad2 deletion exhibited accelerated formation and malignant progression of chemically induced skin tumors compared with WT mice. Consistent with the loss of Smad2 in poorly differentiated human SCCs, Smad2 -/-tumors were poorly differentiated and underwent epithelial-mesenchymal transition (EMT) prior to spontaneous Smad4 loss. Reduced E-cadherin and activation of its transcriptional repressor Snail were also found in Smad2 -/-mouse epidermis and occurred more frequently in Smad2-negative human SCCs than in Smad2-positive SCCs. Knocking down Snail abrogated Smad2 loss-associated EMT, suggesting that Snail upregulation is a major mediator of Smad2 loss-associated EMT. Furthermore, Smad2 loss led to a significant increase in Smad4 binding to the Snail promoter, and knocking down either Smad3 or Smad4 in keratinocytes abrogated Smad2 loss-associated Snail overexpression. Our data suggest that enhanced Smad3/Smad4-mediated Snail transcription contributed to Smad2 loss-associated EMT during skin carcinogenesis.

The functions of transforming growth factor beta-1(TGFbeta1) are cell-context specific. We have found that TGFbeta1 expression in human skin squamous cell carcinoma (SCC) samples has two distinct distribution patterns: (1) either predominantly in suprabasal layers or (2) throughout tumor epithelia including basal proliferative cells. To understand whether the spatial TGFbeta1 expression patterns affect its functions, we have generated several keratinocyte-specific transgenic mouse models in which TGFbeta1 overexpression can be induced either predominantly in the suprabasal epidermis or in the basal layer of the epidermis and hair follicles. Suprabasal TGFbeta1 overexpression inhibits keratinocyte proliferation, suppresses skin carcinogenesis at early stages, but promotes tumor invasion at later stages. In contrast, TGFbeta1 overexpression in the basal layer of the epidermis and hair follicles causes a severe inflammatory skin disorder and epidermal hyperproliferation. Given the importance of inflammation in cancer development, our data suggest that TGFbeta1-induced skin inflammation may override its tumor suppressive effect at early stages during skin carcinogenesis. This hypothesis is further suggested by our recent study that Smad3 knockout mice are resistant to skin chemical carcinogenesis at least in part via abrogation of endogenous TGFbeta1-induced inflammation. This review intends to summarize current insights into the role of TGFbeta1 in skin inflammation and carcinogenesis.

TGF-β signaling can promote tumor formation and development or suppress it, depending on the cellular context and tumor stage. A potential target of this dual effect of TGF-β is HGF, as TGF-β can inhibit or promote its expression, although the mechanisms underlying this are largely unknown. In the present study, we found that mice with keratinocyte-specific deletion of the TGF-β signaling mediator Smad2 (referred to herein as K5.Smad2 -/-mice), which have increased susceptibility to squamous cell carcinomas (SCCs), exhibited angiogenesis associated with epithelial overexpression of HGF and endothelial activation of the HGF receptor c-Met. Application of a c-Met inhibitor abrogated angiogenesis, suggesting that HGF overexpression plays a major role in angiogenesis associated with epithelial Smad2 loss. On the Hgf promoter, Smad2 was mainly associated with transcriptional corepressors, whereas Smad4 was mainly associated with the transcriptional coactivator CREB-binding protein (CBP/p300). Smad2 loss caused increased binding of Smad4 and CBP/p300 to the Hgf promoter. Consistent with this, knocking down Smad2 in human keratinocytes caused increased levels of HGF, which were abrogated by concomitant knockdown of Smad3 and Smad4. Importantly, the incidence of HGF-positive human SCC was high in cases with Smad2 loss and lower when Smad4 was also lost. We therefore conclude that Smad2 loss causes HGF upregulation via loss of Smad2-mediated transcriptional repression and enhanced Smad3/4-mediated transactivation. Since Smad2 is often downregulated in human SCCs, our data suggest a therapeutic strategy of blocking HGF/c-Met activation for Smad2-deficient SCCs.

In the Methods section, the authors inadvertently provided incorrect quantitative PCR primer sequences for amplification of mouse and human HGF. The correct passage appears below.HGF levels were determined using Power SYBR Green Master Mix (Applied Biosystems) and the following custom primers: 5ʹ-AGGAACAGGGGCTTTACGTT-3ʹ (forward) and 5ʹ-GTCAAATTCATGGCCAAACC -3ʹ (reverse). Human HGF quantitative PCR assays were performed using the TaqMan probe set Hs00300159_m1.The authors regret the error.

Transforming growth factor beta (TGFbeta) signaling has both tumor suppression and promotion roles. Smads are transcription factors that primarily mediate intracellular signaling for the TGFbeta superfamily. Loss of Smad2 and Smad4, but not Smad3 is common in human cancers. Given the complex nature of TGFbeta signaling, dissection of the distinct role of each Smad in mediating the multiple functions of TGFbeta signaling is warranted. To further analyze Smad deregulation during carcinogenesis, Smad2, Smad3, Smad4, and Smad7 were genetically modified in murine epidermis, and each alteration resulted in distinct skin phenotypes. Based on data from human cancer samples and from experimental models, Smad2 and Smad4 mainly function as tumor suppressors in skin carcinogenesis in vivo, whereas Smad3 and Smad7 may have dual roles in cancer. This review intends to summarize recent advances in the elucidation of the roles of Smad2, Smad3, Smad4, and Smad7 in skin carcinogenesis.

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