Discovery and Roles of ER-Endolysosomal Contact Sites in Disease.

Inter-organelle membrane contact sites (MCSs) serve as unique microenvironments for the sensing and exchange of cellular metabolites and lipids. Though poorly defined, ER-endolysosomal contact sites are quickly becoming recognized as centers for inter-organelle lipid exchange and metabolic decision-making. Here, we review the discovery and current state of knowledge of ER-endolysosomal MCSs with particular focus on the molecular players that establish and/or utilize these contact sites in metabolism.

Essential N-terminal insertion motif anchors the ESCRT-III filament during MVB vesicle formation.

The endosomal sorting complexes required for transport (ESCRTs) have emerged as key cellular machinery that drive topologically unique membrane deformation and scission. Understanding how the ESCRT-III polymer interacts with membrane, promoting and/or stabilizing membrane deformation, is an important step in elucidating this sculpting mechanism. Using a combination of genetic and biochemical approaches, both in vivo and in vitro, we identify two essential modules required for ESCRT-III-membrane association: an electrostatic cluster and an N-terminal insertion motif.

Molecular mechanisms of the membrane sculpting ESCRT pathway.

The endosomal sorting complexes required for transport (ESCRT) drive multivesicular body (MVB) biogenesis and cytokinetic abscission. Originally identified through genetics and cell biology, more recent work has begun to elucidate the molecular mechanisms of ESCRT-mediated membrane remodeling, with special focus on the ESCRT-III complex. In particular, several light and electron microscopic studies provide high-resolution imaging of ESCRT-III rings and spirals that purportedly drive MVB morphogenesis and abscission.

The endosomal sorting complex ESCRT-II mediates the assembly and architecture of ESCRT-III helices.

The endosomal sorting complexes required for transport (ESCRTs) constitute hetero-oligomeric machines that mediate topologically similar membrane-sculpting processes, including cytokinesis, retroviral egress, and multivesicular body (MVB) biogenesis. Although ESCRT-III drives membrane remodeling that creates MVBs, its structure and the mechanism of vesicle formation are unclear. Using electron microscopy, we visualize an ESCRT-II:ESCRT-III supercomplex and propose how it mediates vesicle formation.

Intracellular curvature-generating proteins in cell-to-cell fusion.

Cell-to-cell fusion plays an important role in normal physiology and in different pathological conditions. Early fusion stages mediated by specialized proteins and yielding fusion pores are followed by a pore expansion stage that is dependent on cell metabolism and yet unidentified machinery. Because of a similarity of membrane bending in the fusion pore rim and in highly curved intracellular membrane compartments, in the present study we explored whether changes in the activity of the proteins that generate these compartments affect cell fusion initiated by protein fusogens of influenza virus and baculovirus.

FCHo proteins are nucleators of clathrin-mediated endocytosis.

Clathrin-mediated endocytosis, the major pathway for ligand internalization into eukaryotic cells, is thought to be initiated by the clustering of clathrin and adaptors around receptors destined for internalization. However, here we report that the membrane-sculpting F-BAR domain-containing Fer/Cip4 homology domain-only proteins 1 and 2 (FCHo1/2) were required for plasma membrane clathrin-coated vesicle (CCV) budding and marked sites of CCV formation. Changes in FCHo1/2 expression levels correlated directly with numbers of CCV budding events, ligand endocytosis, and synaptic vesicle marker recycling.

Structure and analysis of FCHo2 F-BAR domain: a dimerizing and membrane recruitment module that effects membrane curvature.

A spectrum of membrane curvatures exists within cells, and proteins have evolved different modules to detect, create, and maintain these curvatures. Here we present the crystal structure of one such module found within human FCHo2. This F-BAR (extended FCH) module consists of two F-BAR domains, forming an intrinsically curved all-helical antiparallel dimer with a Kd of 2.