A positive FGFR3/FOXN1 feedback loop underlies benign skin keratosis versus squamous cell carcinoma formation in humans
Anna Mandinova, … , Janice L. Brissette, G. Paolo Dotto
Anna Mandinova, … , Janice L. Brissette, G. Paolo Dotto
Published September 1, 2009
Citation Information: J Clin Invest. 2009;119(10):3127-3137. https://doi.org/10.1172/JCI38543.
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Research Article Dermatology

A positive FGFR3/FOXN1 feedback loop underlies benign skin keratosis versus squamous cell carcinoma formation in humans

Abstract

Seborrheic keratoses (SKs) are common, benign epithelial tumors of the skin that do not, or very rarely, progress into malignancy, for reasons that are not understood. We investigated this by gene expression profiling of human SKs and cutaneous squamous cell carcinomas (SCCs) and found that several genes previously connected with keratinocyte tumor development were similarly modulated in SKs and SCCs, whereas the expression of others differed by only a few fold. In contrast, the tyrosine kinase receptor FGF receptor–3 (FGFR3) and the transcription factor forkhead box N1 (FOXN1) were highly expressed in SKs, and close to undetectable in SCCs. We also showed that increased FGFR3 activity was sufficient to induce FOXN1 expression, counteract the inhibitory effect of EGFR signaling on FOXN1 expression and differentiation, and induce differentiation in a FOXN1-dependent manner. Knockdown of FOXN1 expression in primary human keratinocytes cooperated with oncogenic RAS in the induction of SCC-like tumors, whereas increased FOXN1 expression triggered the SCC cells to shift to a benign SK-like tumor phenotype, which included increased FGFR3 expression. Thus, we have uncovered a positive regulatory loop between FGFR3 and FOXN1 that underlies a benign versus malignant skin tumor phenotype.

Authors

Anna Mandinova, Vihren Kolev, Victor Neel, Bing Hu, Wesley Stonely, Jocelyn Lieb, Xunwei Wu, Claudia Colli, Rong Han, Mike Pazin, Paola Ostano, Reinhard Dummer, Janice L. Brissette, G. Paolo Dotto

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Figure 4

Opposite control of FOXN1 expression by EGFR and FGFR3.

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Opposite control of FOXN1 expression by EGFR and FGFR3. (A and B) Human ...
(A and B) Human keratinocytes were transfected with siRNAs for EGFR, ERK1 (A), FGFR3 (B), or control siRNAs, and levels of FOXN1 mRNA were measured by real-time RT-PCR. For internal normalization, here and in all subsequent analyses, 36B4 was used. (C and D) Human keratinocytes were treated with 1 ng/ml EGF or 5 ng/ml FGF9 for 24 hours, followed by real-time RT-PCR analysis (C) or immunoblotting for FOXN1 expression (D). (E) Human keratinocytes (left) and fresh human skin explants (right) were treated with DMSO or with FGFR (SU5402) or EGFR (AG478) inhibitors and analyzed by real-time RT-PCR for FOXN1. (F) Human keratinocytes were transfected with siRNAs for the indicated genes or with control siRNA. Levels of FOXN1 mRNA were assessed by real-time RT-PCR. (G) Human keratinocytes were processed for ChIP with an antibody against c-Jun and control rabbit IgG. Real-time PCR was used to amplify regions of the human FOXN1 promoter using specific primers (Supplemental Table 3). (H) Human keratinocytes were cotransfected with a FOXN1 reporter together with a c-Jun plasmid (c-Jun–CMV; ref. 60) or empty vector control (CMV). Luciferase activity was determined using Renilla for normalization. (I) Human keratinocytes were cotransfected with a FOXN1 reporter with or without expression vectors for wild-type FGFR3, a constitutively active (CA) mutant (61), or control. Promoter activity was measured as described above. (J) Human keratinocytes were treated as in C and D, followed by immunoblot for phosphorylated (p) and total (t) c-Jun and ERK1/2. All error bars denote SEM.