Cell Biology
WNT1-inducible signaling pathway protein 1 (WISP1/CCN4) stimulates melanoma invasion and metastasis by promoting the epithelial–mesenchymal transition

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Besides intrinsic changes, malignant cells also release soluble signals that reshape their microenvironment. Among these signals is WNT1-inducible signaling pathway protein 1 (WISP1), a secreted matricellular protein whose expression is elevated in several cancers, including melanoma, and is associated with reduced survival of patients diagnosed with primary melanoma. Here, we found that WISP1 knockout increases cell proliferation and represses wound healing, migration, and invasion of mouse and human melanoma cells in multiple in vitro assays. Metastasis assays revealed that WISP1 knockout represses tumor metastasis of B16F10 and YUMM1.7 melanoma cells in both C57BL/6Ncrl and NOD-scid IL2Rγnull (NSG) mice. WT B16F10 cells having an invasion phenotype in a transwell assay possessed a gene expression signature similar to that observed in the epithelial–mesenchymal transition (EMT), including E-cadherin repression and fibronectin and N-cadherin induction. Upon WISP1 knockout, expression of these EMT signature genes went in the opposite direction in both mouse and human cell lines, and EMT-associated gene expression was restored upon exposure to media containing WISP1 or to recombinant WISP1 protein. In vivo, Wisp1 knockout–associated metastasis repression was reversed by the reintroduction of either WISP1 or snail family transcriptional repressor 1 (SNAI1). Experiments testing EMT gene activation and inhibition with recombinant WISP1 or kinase inhibitors in B16F10 and YUMM1.7 cells suggested that WISP1 activates AKT Ser/Thr kinase and that MEK/ERK signaling pathways shift melanoma cells from proliferation to invasion. Our results indicate that WISP1 present within the tumor microenvironment stimulates melanoma invasion and metastasis by promoting an EMT-like process.

melanoma
metastasis
invasion
epithelial-mesenchymal transition (EMT)
tumor microenvironment
cancer
Wnt signaling
AKT signaling
secreted protein
WNT1-inducible signaling pathway protein 1 (WISP1/CCN4)

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This work was supported by National Science Foundation Grant CBET-1644932 (to D. J. K.) and NCI, National Institutes of Health, Grant R01CA193473 (to D. J. K.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

This article contains Tables S1–S3 and Figs. S1–S9.

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The abbreviations used are:

    EMT

    epithelial–mesenchymal transition

    EMT-TF

    EMT-inducing transcription factor

    qPCR and qRT-PCR

    quantitative PCR and quantitative RT-PCR, respectively

    TGF

    transforming growth factor

    CCN

    cellular communication network

    MAPK

    mitogen-activated protein kinase

    ERK

    extracellular signal-regulated kinase

    MEK

    mitogen-activated protein kinase/extracellular signal-regulated kinase kinase

    PI3K

    phosphatidylinositol 3-kinase

    JNK

    c-Jun N-terminal kinase

    IHC

    immunohistochemistry

    FPKM

    fragments per kilobase of transcript per million mapped reads

    2D and 3D

    two- and three-dimensional, respectively

    YUMM

    Yale University Mouse Melanoma

    MET

    mesenchymal–epithelial transition

    DMEM

    Dulbecco's modified Eagle's medium

    FBS

    fetal bovine serum

    SFM

    serum-free medium.