HIF1α and HIF2α independently activate SRC to promote melanoma metastases
Sara C. Hanna, Bhavani Krishnan, Sean T. Bailey, Stergios J. Moschos, Pei-Fen Kuan, Takeshi Shimamura, Lukas D. Osborne, Marni B. Siegel, Lyn M. Duncan, E. Tim O’Brien III, Richard Superfine, C. Ryan Miller, M. Celeste Simon, Kwok-Kin Wong, William Y. Kim
Sara C. Hanna, Bhavani Krishnan, Sean T. Bailey, Stergios J. Moschos, Pei-Fen Kuan, Takeshi Shimamura, Lukas D. Osborne, Marni B. Siegel, Lyn M. Duncan, E. Tim O’Brien III, Richard Superfine, C. Ryan Miller, M. Celeste Simon, Kwok-Kin Wong, William Y. Kim
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Research Article Oncology

HIF1α and HIF2α independently activate SRC to promote melanoma metastases

Abstract

Malignant melanoma is characterized by a propensity for early lymphatic and hematogenous spread. The hypoxia-inducible factor (HIF) family of transcription factors is upregulated in melanoma by key oncogenic drivers. HIFs promote the activation of genes involved in cancer initiation, progression, and metastases. Hypoxia has been shown to enhance the invasiveness and metastatic potential of tumor cells by regulating the genes involved in the breakdown of the ECM as well as genes that control motility and adhesion of tumor cells. Using a Pten-deficient, Braf-mutant genetically engineered mouse model of melanoma, we demonstrated that inactivation of HIF1α or HIF2α abrogates metastasis without affecting primary tumor formation. HIF1α and HIF2α drive melanoma invasion and invadopodia formation through PDGFRα and focal adhesion kinase–mediated (FAK-mediated) activation of SRC and by coordinating ECM degradation via MT1-MMP and MMP2 expression. These results establish the importance of HIFs in melanoma progression and demonstrate that HIF1α and HIF2α activate independent transcriptional programs that promote metastasis by coordinately regulating cell invasion and ECM remodeling.

Authors

Sara C. Hanna, Bhavani Krishnan, Sean T. Bailey, Stergios J. Moschos, Pei-Fen Kuan, Takeshi Shimamura, Lukas D. Osborne, Marni B. Siegel, Lyn M. Duncan, E. Tim O’Brien III, Richard Superfine, C. Ryan Miller, M. Celeste Simon, Kwok-Kin Wong, William Y. Kim

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Figure 3

Knockdown of HIF1α or HIF2α reduces the hypoxia-induced invasion and invadopodia formation of melanoma cell lines.

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Knockdown of HIF1α or HIF2α reduces the hypoxia-induced invasion and inv...
(A) A375 SM and WM2664 cells were transfected with siRNAs against HIF1α, HIF2α, or a nonspecific sequence (NS). Whole-cell lysates were immunoblotted with the indicated antibodies. (B) Representative photomicrographs of A375 SM and WM2664 cells transfected with the indicated siRNAs, which have invaded through Matrigel chambers under hypoxia. Original magnification, ×10. (C) Quantification of A375 SM and WM2664 cells transfected with the indicated siRNAs, which have invaded through Matrigel chambers under hypoxia. (D) Representative immunofluorescence images of siRNA-transfected A375 SM cells plated on Alexa Fluor 568–conjugated fibronectin and stained with the indicated antibodies. Original magnification, ×63. Invadopodia were defined as colocalization of cortactin, F-actin (phalloidin), and degradation of Alexa Fluor 568 fibronectin and are indicated with yellow arrowheads. (E) Quantification of the percentage of cells with active invadopodia in siRNA-transfected A375 SM and WM2664 cells. (F) Quantification of the number of visible lung metastases found in mice tail vein injected with A375 SM cells. (G) Gross representative images of the lungs from tail-vein–injected mice. Yellow arrows indicate lung metastases. Error bars show SEM. ***P < 0.0005; **P < 0.005; *P < 0.05.