Pannexin 1 knockdown in metastatic melanoma cells induces cell differentiation into a melanocytic phenotype decreasing tumor size and metastasis in vivo

Abstract

Pannexin 1 (Panx1) is a channel‐forming glycoprotein expressed in different cell types of mammalian skin. We studied the role of Panx1 in melanoma tumorigenesis and metastasis since the mouse melanocyte line CRL‐2770 showed low levels of Panx1 expression, while Panx1 in isogenic melanoma cell lines (B16‐F0, ‐F10 and ‐BL6) was increasingly up‐regulated according to qPCR and Western blot assays. Panx1 shRNA knockdown generated stable BL6 cell lines (Panx1‐KD) that showed a marked increase in melanocyte‐like cell characteristics, including higher melanin production, decreased cell migration and enhanced formation of cellular projections, while compromising Panx1 dye uptake channel function. Western blotting and proteomic analyses using 2D‐SDS‐PAGE gels/mass spectroscopy identified vimentin and beta‐catenin as some of the markers of malignant melanoma that were down‐regulated in Panx1‐KD cells. In vivo, Panx1‐KD cells were implanted on chorioallantoic membranes of day 10 avian embryos. After seven days, the excised Panx1‐KD tumors were significantly smaller than controls and metastasis to the liver was significantly reduced as quantified by mouse‐Alu qPCR of the excised embryonic organs. These data suggests that Panx1 may play a role in skin melanocytes that is potentially dysregulated upon malignant transformation in metastatic melanomas. Supported by the Canadian Institutes of Health Research.