The major histocompatibility complex (MHC) class II mediated signalling increases lipid raft recruitment of adhesion receptors and signal transduction proteins in melanoma cells.

Background: The Major Histocompatibility Complex (MHC) class II molecules are signalling receptors whose engagement leads to the activation of several signalling proteins often enabled by the lipid raft localisation of these molecules. The lipid rafts are specific microdomains of the plasma membrane enriched in sphingolipids and cholesterol implicated in selective protein-protein interactions as well as in the assembly of transient signalling platforms. Indeed, the aggressive metastatic trend of melanoma is associated to the lipid raft recruitment and to the deregulation of receptor and signalling protein functions that promotes the tumour cells detachment from primary tumour and the tissue borders. Therefore, in the aim to understand the molecular mechanisms used by melanomas to the metastatic progression, we studied in MHC class II constitutive expressing melanoma cell lines, the membrane localization of adhesion receptors and signalling proteins. Material and methods: The class II constitutive expressing melanoma cells (A375 and HT144 cell lines) were stimulated with a specific anti-HLA-DR mAb (L243) that mimics the TCR interaction with the class II molecules, for 24h and 48h or left unstimulated. The lipid rafts of stimulated and unstimulated melanoma cells were isolated and analysed by western blot and co-immunoprecipitation as well as through immunofluorescence experiments. Results: In melanoma cells stimulated with the L243 specific anti-HLA-DR antibody, we showed that the HLA-DR mediated signalling increases the lipid raft localisation of class II molecules, Integrin and CAM adhesion receptors as well as of FAK, AKT and STAT3 signalling proteins. Moreover, we reported the lipid rafts association of some adhesion molecules and signalling proteins in stimulated melanoma cells. Furthermore, we identified the HLA-DR mediated signals depending on lipid rafts integrity through the melanoma cells treatment with methyl-?-cyclodextrin (M?CD) that disrupt the lipid raft domains through cholesterol depletion. Conclusions: Therefore, our results suggest a new model in which the HLA-DR stimulation activates a signalling in melanoma cells that increases the lipid raft localization and association between class II molecules, adhesion receptors and signalling molecules, providing a platform useful to frustrate an effective anti-tumour response.