ABSTRACT
Cell-cell contacts inhibit cell growth and proliferation in part by activating the Hippo pathway that drives the phosphorylation and nuclear exclusion of the transcriptional coactivators YAP and TAZ. Cell density and Hippo signaling have also been reported to block transforming growth factor ß (TGF-ß) responses, based on the ability of phospho-YAP/TAZ to sequester TGF-ß-activated SMAD complexes in the cytoplasm. Herein, we provide evidence that epithelial cell polarization interferes with TGF-ß signaling well upstream and independent of cytoplasmic YAP/TAZ. Rather, polarized basolateral presentation of TGF-ß receptors I and II deprives apically delivered TGF-ß of access to its receptors. Basolateral ligand delivery nonetheless remains entirely effective to induce TGF-ß responses. These data demonstrate that cell-type-specific inhibition of TGF-ß signaling by cell density is restricted to polarized epithelial cells and reflects the polarized distribution of TGF-ß receptors, which thus affects SMAD activation irrespective of Hippo pathway activation.
Subject(s)
Cytoplasm/metabolism , Nuclear Proteins/metabolism , Protein Serine-Threonine Kinases/metabolism , Receptors, Transforming Growth Factor beta/metabolism , Signal Transduction , Transcription Factors/metabolism , Transforming Growth Factor beta/metabolism , Acyltransferases , Blotting, Western , Cell Count , Cell Cycle Proteins , Cell Proliferation , Cells, Cultured , Fluorescent Antibody Technique , Hippo Signaling Pathway , Humans , Immunoenzyme Techniques , Immunoprecipitation , Nuclear Proteins/genetics , Phosphorylation , Protein Serine-Threonine Kinases/genetics , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Transcription Factors/genetics , Transforming Growth Factor beta/geneticsABSTRACT
YAP and its paralog protein TAZ are downstream effectors of the Hippo pathway. Both are amplified in many human cancers and promote cell proliferation and epithelial-mesenchymal transition. Little is known about the status of the Hippo pathway in cutaneous melanoma. We profiled Hippo pathway component expression in a panel of human melanoma cell lines and melanocytic lesions, and characterized the capacity of YAP and TAZ to control melanoma cell behavior. YAP and TAZ immuno-staining in human samples revealed mixed cytoplasmic and nuclear staining for both proteins in benign nevi and superficial spreading melanoma. TAZ was expressed at higher levels than YAP1/2 in all cell lines and in those with high invasive potential. Stable YAP or TAZ knockdown dramatically reduced the expression of the classical Hippo target CCN2/connective-tissue growth factor (CTGF), as well as anchorage-independent growth, capacity to invade Matrigel, and ability form lung metastases in mice following tail-vein injection. YAP knockdown also reduced invasion in a model of skin reconstruct. Inversely, YAP overexpression increased melanoma cell invasiveness, associated with increased TEA domain-dependent transcription and CCN2/CTGF expression. Together, these results demonstrate that both YAP and TAZ contribute to the invasive and metastatic capacity of melanoma cells and may represent worthy targets for therapeutic intervention.
