Mechanisms and Intracellular Components in Exocytosis, Membrane Assembly and Repair, and Parasite Replication

Eunice Kennedy Shriver National Institute Of Child Health & Human Development

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Abstract

Living cells are open non-equilibrium systems. To exist, a cell requires precisely controlled maintenance of gradients in the chemical potential, between the extracellular environment, the cytoplasm, and the lumen of organelles, of many constituents. The amphiphilic nature of lipid molecules, self-assembling into lipid bilayers, provides an extremely low permeability barrier to both electrolytes and large nonelectrolytes. For the processes of life, a continuous exchange of matter must occur across all membranes. For example, uptake of large molecules and compounds from the outside occurs via endocytosis, phagocytosis, and macro and micro pinocytosis, secretion occurs via exocytosis, and intracellular protein trafficking via transport vesicles between endoplasmic reticulum, Golgi apparatus, endosomes, and lysosomes. The movement of such membrane-bound cargo dictates membrane recycling, or cells and organelles would be incapable of maintaining their volumes and shapes. These ubiquitous and multifarious events, plus the accommodation of lipid bilayers to membrane proteins and transient pores, all require the lipid bilayer to change its topology. Physical models of these topological changes, essential to life, must take into account the energy of membrane deformations within the framework of an adequate theory of elasticity. All of these biophysical parameters are equally important for communication between cells by extracellular vesicles. Thin sections of HUVECs revealed clusters of membrane protrusions on the otherwise smooth cell surface. The protrusions contained membrane-bound organelles, including multivesicular bodies (MVBs), and appeared to be on the verge of pinching off to form microvesicles. Beyond cell peripheries, membrane-bound vesicles with internal MVBs were observed, and serial sections confirmed that they were not connected to cells. These observations are consistent with the notion that these multi-compartmented microvesicles (MCMVs) pinch-off from protrusions. Remarkably, omega figures formed by fusion of vesicles with the MCMV limiting membrane were directly observed, apparently releasing exosomes from the MCMV. In summary, MCMVs are a novel form of EV that bud from membrane protrusions on the HUVEC surface, contain MVBs and release exosomes. These observations suggest that exosomes can be harboured within and released from transiting microvesicles after departure from the parent cell, constituting a new site of exosome biogenesis occurring from endothelial and potentially additional cell types.