PKC and PKA phosphorylation affect the subcellular localization of claudin-1 in melanoma cells.

Cytoplasmic expression of claudin-1 in metastatic melanoma cells correlates to increased migration, and increased secretion of MMP-2 in a PKC dependent manner, whereas claudin-1 nuclear expression is found in benign nevi. Melanoma cells were transfected with a vector expressing CLDN-1 fused to a nuclear localization signal (NLS). Despite significant nuclear localization of claudin-1, there was still transport of claudin-1 to the cytoplasm.

Assaying Wnt5A-mediated invasion in melanoma cells.

Wnt5A has been implicated in melanoma metastasis, and the progression of other cancers including pancreatic, gastric, prostate, and lung cancers. Assays to test motility and invasion include both in vivo assays and in vitro assays. The in vivo assays include the use of tail vein or footpad injections of metastatic cells, and are often laborious and expensive.

Wnt5A regulates expression of tumor-associated antigens in melanoma via changes in signal transducers and activators of transcription 3 phosphorylation.

There are currently no effective therapies for metastatic melanoma and targeted immunotherapy results in the remission of only a very small percentage of tumors. In this study, we show that the noncanonical Wnt ligand, Wnt5A, can increase melanoma metastasis in vivo while down-regulating the expression of tumor-associated antigens important in eliciting CTL responses (e.g.

The Wnt5A/protein kinase C pathway mediates motility in melanoma cells via the inhibition of metastasis suppressors and initiation of an epithelial to mesenchymal transition.

We have shown that Wnt5A increases the motility of melanoma cells. To explore cellular pathways involving Wnt5A, we compared gain-of-function (WNT5A stable transfectants) versus loss-of-function (siRNA knockdown) of WNT5A by microarray analysis. Increasing WNT5A suppressed the expression of several genes, which were re-expressed after small interference RNA-mediated knockdown of WNT5A.

Claudin-1 overexpression in melanoma is regulated by PKC and contributes to melanoma cell motility.

Serial analysis of gene expression followed by pathway analysis implicated the tight junction protein claudin-1 (CLDN1) in melanoma progression. Tight junction proteins regulate the paracellular transport of molecules, but staining of a tissue microarray revealed that claudin-1 was overexpressed in melanoma, and aberrantly expressed in the cytoplasm of malignant cells, suggesting a role other than transport. Indeed, melanoma cells in culture demonstrate no tight junction function.