Introduction: The Rab GTPase family has been increasingly related to carcinogenesis and cancer biology during the past decade. In melanoma, Rab27a has been identified as a tumor dependency gene. Rab27a is thought to regulate exosome secretion in several cell types, however the exact role of Rab27a in melanoma biology and the underlying mechanisms are not well investigated.
Experimental Procedures: To investigate the influence of Rab27a on melanoma biology and exosome secretion, Rab27a shRNA knockdown (KD) or CRISPR knockout (KO) was performed in human and murine melanoma cell lines. The effect of Rab27a loss on melanoma cell invasion and motility was analyzed by in vitro 3D spheroid invasion and live cell imaging. Rab27a function on melanoma metastasis was investigated using a mouse melanoma spontaneous metastasis model. Exosomes secreted by melanoma cell lines with/without Rab27a KD/KO were purified and characterized by electron microscopy, NanoSight analysis, BCA assay, western blotting and mass spectrometry.
Results: Our results indicate that Rab27a loss in Rab27a high metastatic melanoma cell lines reduced 3D spheroid invasion and cell motility and Rab27a loss also reduced spontaneous melanoma metastasis in vivo. Rab27a KD invasion phenotype can be partially rescued by addition of exosomes from Rab27a replete cell conditioned media, but not exosomes derived from Rab27a KD cells. Loss of Rab27a does not alter the number of exosomes secreted from melanoma cells, but does alter exosome protein composition and morphology. Mass spectrometry analysis has shown that Rab27a replete cell secreted exosomes are more involved in regulating cell invasion and motility than the exosomes from Rab27a KD cells.
Conclusion: In summary, Rab27a promotes the invasion and metastasis of a subset of melanoma cells via the regulation of pro-invasive exosomes, which indicates Rab27a as a potential therapeutic target for preventing melanoma progression.
Citation Format: DAJIANG GUO, Goldie Y. Lui, Danae M. Sharp, Siew Li Lai, Shweta Tikoo, Rain Kwan, Michelle van Geldermalsen, Jeff Holst, Nikolas K. Haass, Wolfgang Weninger, Kimberley A. Beaumont. Rab27a promotes melanoma cell invasion and metastasis via the regulation of exosome secretion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5167.