Streptomyces coelicolor Vesicles: Many Molecules To Be Delivered.

Streptomyces coelicolor is a model organism for the study of Streptomyces, a genus of Gram-positive bacteria that undergoes a complex life cycle and produces a broad repertoire of bioactive metabolites and extracellular enzymes. This study investigated the production and characterization of membrane vesicles (MVs) in liquid cultures of S. coelicolor M145 from a structural and biochemical point of view; this was achieved by combining microscopic, physical and -omics analyses. Two main populations of MVs, with different sizes and cargos, were isolated and purified. S. coelicolor MV cargo was determined to be complex, containing different kinds of proteins and metabolites. In particular, a total of 166 proteins involved in cell metabolism/differentiation, molecular processing/transport, and stress response were identified in MVs, the latter functional class also being important for bacterial morpho-physiological differentiation. A subset of these proteins was protected from degradation following treatment of MVs with proteinase K, indicating their localization inside the vesicles. Moreover, S. coelicolor MVs contained an array of metabolites, such as antibiotics, vitamins, amino acids, and components of carbon metabolism. In conclusion, this analysis provides detailed information on S. coelicolor MVs under basal conditions and on their corresponding content, which may be useful in the near future to elucidate vesicle biogenesis and functions. IMPORTANCE Streptomycetes are widely distributed in nature and characterized by a complex life cycle that involves morphological differentiation. They are very relevant in industry because they produce about half of all clinically used antibiotics, as well as other important pharmaceutical products of natural origin. Streptomyces coelicolor is a model organism for the study of bacterial differentiation and bioactive molecule production. S. coelicolor produces extracellular vesicles that carry many molecules, such as proteins and metabolites, including antibiotics. The elucidation of S. coelicolor extracellular vesicle cargo will help us to understand different aspects of streptomycete physiology, such as cell communication during differentiation and response to environmental stimuli. Moreover, the capability of these vesicles for carrying different kinds of biomolecules opens up new biotechnological possibilities related to drug delivery. Indeed, decoding the molecular mechanisms involved in cargo selection may lead to the customization of extracellular vesicle content.

Effects of human leukocyte antigen (HLA)-DR engagement on melanoma cells.

Melanoma cells often display constitutive expression of the major histocompatibility complex (MHC) class II molecules which is associated with a higher metastatic dissemination. The MHC class II molecules during T cell/ professional antigen-presenting cell (APC) interactions are localized, as signalling receptors, into membrane lipid rafts which are thought to be sites of signalling complex assembly. Therefore, with the aim of understanding the molecular mechanisms used by melanoma cells to frustrate an effective anti-tumour response we stimulated a MHC class II constitutive expressing melanoma cell line with a specific antibody that mimics the interaction of T-cell receptor (TCR) with class II molecules. In stimulated melanoma cells we showed through Western blotting and immunofluorescence experiments the recruitment of HLA-DR molecules in lipid raft compartments as well as the c-Jun NH2-terminal kinase activations as a consequence of the class II engagement. Furthermore, we showed that SDS-stable HLA-DR-peptide complexes are recruited in lipid rafts of stimulated melanoma cells. Therefore, in light of the results reported, our hypothesis is that the redistribution of class II molecules into lipid raft microdomains of stimulated melanoma cells as well as the associated activation of signalling pathways, could be useful for melanoma cells to frustrate an effective anti-tumour response.