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 2

Accelerated tumor formation and malignant conversion of skin carcinogenesis in K5.Smad2-knockout mice.

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Accelerated tumor formation and malignant conversion of skin carcinogene...
(A) Kinetics of tumor formation. Arrow indicates TPA withdrawal. The seemingly more rapid tumor regression after TPA withdrawal in Smad2+/– and Smad2–/– groups compared with Smad2+/+ is due to necessity of euthanizing mice with a higher tumor burden. P < 0.001 compared with K5.Smad2–/– or K5.Smad2+/– and Smad2+/+ (B) Kinetics of malignant conversion. Smad2+/– and Smad2–/– mice had higher rates of malignant conversion (P < 0.05 compared with Smad2+/+ mice). The total number of mice of each genotype was used as a denominator for all time points through the entire course of tumor kinetics in A and B. (C) Tumor pathology and keratin markers. H&E staining of K5.Smad2–/– tumors showed less differentiation compared with K5.Smad2+/+ tumors. Immunofluorescence staining revealed that at the same histological stage, Smad2+/+ papillomas (Paps) expressed K1 (green) in suprabasal layers, whereas K5.Smad2–/– papillomas expressed K8 (red) and almost lost K1 in suprabasal layers. The dotted lines delineate the basement membrane. At SCC stages, K5.Smad2–/– tumors developed spindle cell carcinomas (SPCCs) when K5.Smad2+/+ tumors were well-differentiated SCCs. Rectangles in the bottom 2 panels denote areas of transition from SCC to SPCC. Two of these regions are enlarged 4 times to illustrate this transition (top row, far right panels). Scale bars: 100 μm.