Biochemical and biological characterization of exosomes containing prominin-1/CD133.

ABSTRACT: Exosomes can be viewed as complex "messages" packaged to survive trips to other cells in the local microenvironment and, through body fluids, to distant sites. A large body of evidence indicates a pro-metastatic role for certain types of cancer exosomes. We previously reported that prominin-1 had a pro-metastatic role in melanoma cells and that microvesicles released from metastatic melanoma cells expressed high levels of prominin-1. With the goal to explore the mechanisms that govern proteo-lipidic-microRNA sorting in cancer exosomes and their potential contribution(s) to the metastatic phenotype, we here employed prominin-1-based immunomagnetic separation in combination with filtration and ultracentrifugation to purify prominin-1-expressing exosomes (prom1-exo) from melanoma and colon carcinoma cells. Prom1-exo contained 154 proteins, including all of the 14 proteins most frequently expressed in exosomes, and multiple pro-metastatic proteins, including CD44, MAPK4K, GTP-binding proteins, ADAM10 and Annexin A2. Their lipid composition resembled that of raft microdomains, with a great enrichment in lyso-phosphatidylcholine, lyso-phosphatidyl-ethanolamine and sphingomyelin. The abundance of tetraspanins and of tetraspanin-associated proteins, together with the high levels of sphingomyelin, suggests that proteolipidic assemblies, probably tetraspanin webs, might be the essential structural determinant in the release process of prominin-1 of stem and cancer stem cells. Micro-RNA profiling revealed 49 species of micro-RNA present at higher concentrations in prom1-exo than in parental cells, including 20 with cancer-related function. Extensive accumulation of prom1-exo was observed 3 h after their addition to cultures of melanoma and bone marrow-derived stromal cells (MSC). Short-term co-culture of melanoma cells and MSC resulted in heterologous prominin-1 transfer. Exposure of MSC to prom1-exo increased their invasiveness. Our study supports the concept that specific populations of cancer exosomes contain multiple determinants of the metastatic potential of the cells from which they are derived.

SUBMITTER: Rappa G 

PROVIDER: S-EPMC3698112 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature