ABSTRACT
The endosomal pathway is responsible for plasma membrane cargo uptake, sorting, and, in many cases, lysosome targeting. Endosome maturation is complex, requiring proper spatiotemporal recruitment of factors that regulate the size, maturity, and positioning of endosomal compartments. In animal cells, it also requires trafficking of endosomes on microtubules. Recent work has revealed the presence of contact sites between some endosomes and the endoplasmic reticulum (ER). Although these contact sites are believed to have multiple functions, the frequency, dynamics, and physical attributes of these contacts are poorly understood. Here we use high-resolution three-dimensional electron microscopy to reveal that ER tubules wrap around endosomes and find that both organelles contact microtubules at or near membrane contact sites. As endosomes traffic, they remain bound to the ER, which causes the tubular ER to rearrange its structure around dynamic endosomes at contact sites. Finally, as endosomes transition through steps of maturation, they become more tightly associated with the ER. The major implication of these results is that endosomes mature and traffic while coupled to the ER membrane rather than in isolation.
Subject(s)
Endoplasmic Reticulum/metabolism , Endosomes/metabolism , Microtubules/metabolism , Animals , COS Cells , Cell Line, Tumor , Chlorocebus aethiops , Endoplasmic Reticulum/ultrastructure , Endosomes/ultrastructure , Humans , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Membrane Proteins/genetics , Membrane Proteins/metabolism , Microscopy, Confocal , Microscopy, Electron , Microtubules/ultrastructure , Multivesicular Bodies/metabolism , Multivesicular Bodies/ultrastructure , Nocodazole/pharmacology , Protein Transport/drug effects , SEC Translocation Channels , Time-Lapse Imaging , rab GTP-Binding Proteins/genetics , rab GTP-Binding Proteins/metabolism , rab5 GTP-Binding Proteins/genetics , rab5 GTP-Binding Proteins/metabolism , rab7 GTP-Binding ProteinsABSTRACT
Mitochondrial structure and distribution are regulated by division and fusion events. Mitochondrial division is regulated by Dnm1/Drp1, a dynamin-related protein that forms helices around mitochondria to mediate fission. Little is known about what determines sites of mitochondrial fission within the mitochondrial network. The endoplasmic reticulum (ER) and mitochondria exhibit tightly coupled dynamics and have extensive contacts. We tested whether ER plays a role in mitochondrial division. We found that mitochondrial division occurred at positions where ER tubules contacted mitochondria and mediated constriction before Drp1 recruitment. Thus, ER tubules may play an active role in defining the position of mitochondrial division sites.
