Table_2_ATG5 and ATG7 Expression Levels Are Reduced in Cutaneous Melanoma and Regulated by NRF1.docx

Autophagy is a highly conserved cellular process in which intracellular proteins and organelles are sequestered and degraded after the fusion of double-membrane vesicles known as autophagosomes with lysosomes. The process of autophagy is dependent on autophagy-related (ATG) proteins. The role of autophagy in cancer is very complex and still elusive. We investigated the expression of ATG proteins in benign nevi, primary and metastatic melanoma tissues using customized tissue microarrays (TMA). Results from immunohistochemistry show that the expression of ATG5 and ATG7 is significantly reduced in melanoma tissues compared to benign nevi. This reduction correlated with changes in the expression of autophagic activity markers, suggesting decreased basal levels of autophagy in primary and metastatic melanomas. Furthermore, the analysis of survival data of melanoma patients revealed an association between reduced ATG5 and ATG7 levels with an unfavourable clinical outcome. Currently, the mechanisms regulating ATG expression levels in human melanoma remains unknown. Using bioinformatic predictions of transcription factor (TF) binding motifs in accessible chromatin of primary melanocytes, we identified new TFs involved in the regulation of core ATGs. We then show that nuclear respiratory factor 1 (NRF1) stimulates the production of mRNA and protein as well as the promoter activity of ATG5 and ATG7. Moreover, NRF1 deficiency increased in vitro migration of melanoma cells. Our results support the concept that reduced autophagic activity contributes to melanoma development and progression, and identifies NRF1 as a novel TF involved in the regulation of both ATG5 and ATG7 genes.