The ubiquitin-specific protease 8 antagonizes melatonin-induced endocytic degradation of MT1 receptor to promote lung adenocarcinoma growth

[Display omitted] •Melatonin can induce the downregulation of the MT1 receptor.•MT1 receptor internalization incurred by melatonin follows the canonical endolysosomal pathway.•The ubiquitin-specific protease 8 antagonizes the endocytic degradation of the MT1 receptor.•The suppression of ubiquitin-specific protease 8 potentiates the cancer-inhibitory effects of melatonin in vitro.•Combination of USP8 inhibition and melatonin treatment effectively deters tumor growth in xenograft mouse models. The human genome encodes two melatonin receptors (MT1 and MT2) that relay melatonin signals to cellular interior. Accumulating evidence has linked melatonin to multiple health benefits, among which its anticancer effects have become well-established. However, the implications of its receptors in lung adenocarcinoma have so far remained incompletely understood. This study aims to investigate the response of the MT1 receptor to melatonin treatment and its dynamic regulation by ubiquitin-specific protease 8 (USP8) in lung adenocarcinoma. The mRNA levels of MT1 and MT2 receptors were analyzed with sequencing data. The expression and localization of the MT1 receptor with melatonin treatment were investigated by immunoblotting, immunofluorescence and confocal microscopy assays. Endocytic deubiquitylases were screened to identify MT1 association. The effects of USP8 were assessed with shRNA-mediated knockdown and small molecule inhibitor. The combined efficacy of melatonin and USP8 suppression was also evaluated using xenograft animal models. Bioinformatic analysis revealed increased expression of the MT1 receptor in lung adenocarcinoma tissues. Melatonin treatment leads to the downregulation of the MT1 receptor in lung adenocarcinoma cells, which is attributed to receptor endocytosis and lysosomal degradation via the canonical endo-lysosomal route. USP8 negatively regulates the endocytic degradation of the MT1 receptor incurred by melatonin exposure and thus protects lung adenocarcinoma cell growth. USP8 suppression by knockdown or pharmacological inhibition effectively deters cancer cell proliferation and sensitizes lung adenocarcinoma cells to melatonin in vitro. Furthermore, USP8 silencing significantly potentiates the anticancer effects of melatonin in xenograft tumor models. The MT1 receptor responds to melatonin treatment and is endocytosed for lysosomal degradation that is counteracted by USP8. The inhibition of USP8 demonstrates tumor-suppressive effects and thus can