Regulation of gap junctions in melanoma and their impact on Melan-A/MART-1-specific CD8+ T lymphocyte emergence
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Gap junctions (GJs) enable intercellular communication between adjacent cells through channels of connexins. Using a three-dimensional construct, we previously showed that endothelial and tumor cells formed GJs, allowing melanoma-specific T lymphocytes to recognize and kill melanoma-derived endothelial cells. We demonstrate here on histological sections of melanoma biopsies that GJ formation occurs in vivo between tumor and endothelial cells and between T lymphocytes and target cells. We also show an in vitro increase of GJ formation in melanoma and endothelial cells following dacarbazin and interferon gamma (IFN-γ) treatment or hypoxic stress induction. Our data indicate that although connexin 43 (Cx43), the main GJ protein of the immune system, was localized at the immunological synapse between T lymphocyte and autologous melanoma cells, its over-expression or inhibition of GJs does not interfere with cytotoxic T lymphocyte (CTL) clone lytic function. In contrast, we showed that inhibition of GJs by oleamide during stimulation of resting PBMCs with Melan-A natural and analog peptides resulted in a decrease in antigen (Ag) specific CD8+ T lymphocyte induction. These Ag-specific CD8+ cells displayed paradoxically stronger reactivity as revealed by CD107a degranulation and IFN-γ secretion. These findings indicate that Cx43 does not affect lytic function of differentiated CTL, but reveal a major role for GJs in the regulation of antigen CD8+-naïve T lymphocyte activation.
GJ formation occurs in vivo between T lymphocytes and tumor cells
Cx43 localized at the immunological synapse between T and autologous melanoma cells
Inhibition of GJs resulted in a decrease in Ag-specific CD8+ T lymphocyte induction
A role for GJs in the regulation of antigen CD8+-naïve T lymphocyte activation
KeywordsMelanoma Cytotoxic T cell Antigen presentation Gap junction Connexin 43
We thank Nadine Gervois and Nathalie Labarrière for the HLA-A2/Melan-A tetramer gift as well as Lisa Bain for manuscript editing.
Cx43 localizes at the immunological synapse. Intracellular trafficking of Cx43-GFP to the contact area between Cx43-GFP transfected M4T and autologous CTL was analyzed by confocal microscopy (AVI 5,082 kb)
Cx43 localizes at the immunological synapse and gives rise to functional GJs. Calcein was transferred from CTL to M4T cells after contact as shown by confocal microscopy (AVI 66,204 kb)
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