Keratinocyte-specific Smad2 ablation results in increased epithelial-mesenchymal transition during skin cancer formation and progression
Kristina E. Hoot, … , Erwin Bottinger, Xiao-Jing Wang
Kristina E. Hoot, … , Erwin Bottinger, Xiao-Jing Wang
Published July 10, 2008
Citation Information: J Clin Invest. 2008;118(8):2722-2732. https://doi.org/10.1172/JCI33713.
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Research Article Oncology

Keratinocyte-specific Smad2 ablation results in increased epithelial-mesenchymal transition during skin cancer formation and progression

Abstract

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.

Authors

Kristina E. Hoot, Jessyka Lighthall, Gangwen Han, Shi-Long Lu, Allen Li, Wenjun Ju, Molly Kulesz-Martin, Erwin Bottinger, Xiao-Jing Wang

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Figure 7

Increased Smad3/Smad4-mediated Snail transcription contributes to Smad2 loss–associated Snail overexpression.

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Increased Smad3/Smad4-mediated Snail transcription contributes to Smad2 ...
(A and B) Comparative PCR from chromatin immunoprecipitation (IP) showed an increase in Smad4 binding to the Snail promoter in Smad2–/– skin compared with WT skin. Residue Smad2 binding in Smad2–/– skin was from nonkeratinocyte population of the whole skin. Smad3 binding to the Snail promoter was not significantly changed in Smad2–/– skin in comparison with WT skin. Smad4 binding to the Snail promoter was significantly increased in Smad2–/– skin. *P < 0.05. (C) Dual knockdown of Smad2 and Smad3 or Smad2 and Smad4 abrogated Smad2 loss–associated Snail overexpression. Smad2 knockdown (48 hours) caused a significant increase in Snail expression. Knockdown of Smad4 alone caused a reduction in Snail expression. Concomitant knockdown of Smad2 and Smad3 or Smad2 and Smad4 reduced Snail expression back to mock-transfection levels. †P < 0.05 compared with mock transfection. ‡P < 0.05 compared with Smad2 siRNA treatment.