Cellular Package Logistics

Some new insights on how vesicle fusion occurs with target membranes.

Membranes were one of the founding inventions of life. Every cell has a membrane, and many viruses do as well. Whether they were present from the start, or were a later innovation when the nascent chemical reactions of life, begun in some rocky pore, freed themselves into the open ocean, the fatty bilayer membrane is now fundamental. And from a proper perspective, it is a formidable barrier. If you were a water molecule, a typical membrane would be a sixty-five foot wide mass of repellent goo.

A typical cellular membrane, with the fatty bilayer core and a few proteins and other molecules sprinkled about, all of which move freely in the plane of the membrane.

Eukaryotes, as usual, took this innovation to whole new level, developing a variety of internal membranes and organelles that were entirely unknown to their bacterial forebears. Foremost of which is the mitochondrion- the symbiotic bacterium that turned into a powerful organelle. But there are many others, like the endoplasmic reticulum, the nucleus, the lysosome, phagosome, and the golgi apparatus. There is constant traffic among these organelles, with small membrane vesicles being emitted, traveling around, and finding and then fusing with their target destination membranes. It is like a tiny FedEx system within each cell, complete with addresses, carriers, cargoes, and grateful recipients.

A cartoon about the some of the internal membranes of eukaryotic cells. Note all the traffic going about. The lysosome, for instance, receives incoming vesicles from the plasma membrane, as things to digest, and from the golgi apparatus, as packages of new enzymes to do its work- enzymes that only turn on in the acidic environment inside the lysosome. This traffic requires a great deal of vesicle formation, transportation and fusion with targets, whose molecular detail is being gradually revealed. The golgi apparatus is a central sorting and distribution center.

Vesicles can exist because they are generally not prone to fuse with each other. Each membrane has a electrical charge-rich exterior that keeps it happily hydrated and slightly stand-offish vs other membranes. So something extra is needed to provide the push to fuse with a target membrane. And this barrier also provides the possibility of regulation, getting the right vesicle to fuse with the right target. A recent paper discussed one small part of this quite complicated process- the structure of tethering proteins that bring cargo vesicles and target membranes together. The research group focused on vesicles destined for the lysosome. The lysosome is the target of two major types of vesicles. One type brings in the enzymes needed for the lysosome to digest food for the cell. The other type are endosomes coming from the plasma membrane, and other sources of cellular garbage, which go to the lysosome to be digested, much like food is digested in our stomachs.

A schematic of the SNARE proteins that operate at the core of membrane fusion. One (here, synaptobrevin) is attached to the cargo vesicle. Another (here, syntaxin) is attached to the target membrane. A third, SNAP-25, supplies two more alpha helices to the 4-helix structure that winds up and brings the two membranes ever closer. The extra protein (synaptotagmin) is a regulator, in this case at the neuronal synapse, which directs fusion to happen in response to electrical neural activation, thereby helping secrete a neurotransmitter and thus propagate the neural signal.

Vesicle fusion is, at core, conducted by proteins called SNARE proteins. One extends from the cargo vesicle, another extends from the target membrane, and a third joins in, complexed with the second. Their alpha helical structures strongly and progressively inter-twine together to drive the membranes together, forcing fusion. Energetic studies show that at least three of these complexes are needed to get two membranes fused. After the fusion event, the SNARE proteins are recovered by special chaperone proteins that expend ATP to unwind the SNARE helices and reset them for another round of action.

Part of the regulation of this process (of which a great deal remains unknown) is provided by "tethering" complexes- proteins that grab hold of membranes of the right sort, extend across the gap to the target membrane, and also bind the SNARE proteins to orchestrate the fusion process. This research group studied one such tethering complex called HOPS, from yeast cells, which is composed of six proteins, Vps11, 16, 18, 39, 41, fall named for vacuole protein sorting. In yeast cells, the lysosome is called the vacuole, and these were all picked up by genetic screens for defects in getting proteins to the vacuole. Thus HOPS is essential for the fusion process, even though it plays a helping, orchestrating role.

New structures of the HOPS complex, which helps direct cargo vesicles to lysosomes. It is composed of six Vps proteins, all similar, which reach in three directions- to each membrane, and to the SNARE proteins as well. A three-handed helper, as it were.

The researchers found that the HOPS complex is fundamentally a triangle. Two of the ends (green, above) extend to the membranes to be fused, while the third end (brown) engages the SNARE proteins and helps them do their thing when the right geometry has been attained. The shape almost tells the story of what is going on, with the SNARE proteins sitting right in the middle, at the presumptive cleft between the two membranes. The structure has an interesting profusion of beta-propeller structures on all its ends. These are bulky protein domains very commonly used for protein-protein interactions. The shared structures also show that these proteins are deeply related to each other, probably all evolved from a single ancestor. 

Model of HOPS function, as it joins the two membranes, and also orchestrates SNARE action.

The HOPS complex tethers to Ypt7 proteins on both membranes. But Ypt7 is itself highly regulated, and not always "on", i.e. receptive for docking. It is turned on by other proteins that specify that it is in the right place and near the right partner to activate. But that is another story, and one still developing.

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