Loss of p14 reduces immunogenicity in melanoma by affecting the peptide surface density via non-canonical Wnt signaling
Immunotherapy has achieved tremendous success in melanoma. However, only around 50 % of advanced melanoma patients benefit from immunotherapy with immunogenicity of melanomas playing a key role. CDKN2A encoding the two tumor-suppressor proteins p14ARF and p16INK4a belongs to the most frequently inactivated gene loci in advanced melanoma where decreased T cell infiltration is accompanied. While the role of p16INK4a has been aberrantly investigated, knowledge about the function of p14ARF in melanoma therapy is scarce. In this study, we elucidate the impact of reduced p14ARF expression on melanoma immunogenicity. Knockdown of p14ARF in melanoma cell lines diminished the recognition and killing by melanoma differentiation antigen (MDA)-specific T cells. Resistance was caused by a reduction of the peptide surface density of presented MDAs. While antigen presentation via HLA-I molecules was enhanced upon p14ARF downregulation in general, absolute and relative expression of cognate peptides was decreased. Limiting WNT5A signaling reverted this phenotype suggesting an involvement of non-canonical Wnt signaling. Taken together, our data indicate a new mechanism limiting MDA-specific T cell responses by decreasing both absolute and relative MDA-peptide presentation in melanoma. Overall design: To investigate the influence of p14 and p16, melanoma cell line MaMel51 was knocked down for p14, p16 or both of them with respective doxycyclin-inducible shRNAs or a control scramble (scr) shRNA. Three biological triplicates were analyzed for each condition.