A PI3K-WIPI2 loop activates LC3 lipidation in autophagy

This Stop Motion Animation shows the events that take place in the human cell during a process that scientists call Autophagy (from Greek auto- = self and -phagy = eating), in which surplus material inside the cell is removed by cellular digestion. As a consequence of their normal activity, cells produce waste on the daily basis and a disposal mechanism arose during evolution that allows them to recycle and keep their environment clean.
During autophagy a new organelle forms, named autophagosome, which looks and functions just like a rubbish bag. Its role is to wrap the waste and deliver it to the compartment of the cell specialised in degradation: the lysosome, a waste burning station. To this end, the cell first has to build the bag and then fill it with the garbage. This video shows how cells produce autophagic bags.
Generally, bags in cells are made out of lipids, oily molecules that arrange into thin layers called membranes. They divide wet compartments from each other and avoid them from mixing, just like oil and water do when you try to mingle them in your kitchen: they never dissolve! In the laboratory, scientists can prepare membranes called Giant Unilamellar Vesicles (GUVs, as they are pretty big when compared to the cellular scales) and can use them to mimic bags.
But what makes a membrane the right bag for autophagy? All autophagic membranes are marked with a tag, a small molecule called LC3 that functions as the instruction lines on recycling bags that say ‘Paper’ or ‘Metal’. LC3 is attached with a strong molecular glue to a lipid in the membrane called Phosphatidylethanolamine, or PE. This lipid can also be used to prepare GUVs in the laboratory. But that’s not enough..
As not all the PE-containing membranes in cells can become autophagic, other events must happen that kick off the formation of an autophagosome at a given time and in the right place. An actual construction site must be organised and built on these membranes, that involves the work of many different molecular machines.
At first, a signalling lipid must be generated. This lipid is called Phosphatidyl-Inositol(3)Phosphate, or PI(3)P. Nature employs a specialised machine called PI3K complex that can instantly produce PI(3)P, composed of four other subunits: Beclin1, ATG14, VPS34 and VPS15 that all work together for the same purpose. They produce PI(3)P using Adenosine Three Phosphate (or ATP), a coin used in the cell for pricy transactions.
Once this signal is on membranes, it triggers the recruitment of other machines. The next one to arrive is WIPI2. WIPI2 has high affinity for PI(3)P, and as it happens with iron for magnets, it is attracted to membranes rich in PI(3)P and sticks to them. C. Chang discovered that when WIPI2 is around, the PI3K complex can work much better and WIPI2 makes its work more efficient: the more PI(3)P is produced, the more WIPI2 molecules are recruited, thus better the PI3K complex works. So, as it happens in real life, when more workers do the job, the construction site proceeds faster. That’s how nature optimises the job, too!
In the second part, WIPI2 recruits another big complex machine made of three subunits called ATG12–ATG5/ATG16L1, also known altogether as E3. As the name suggests, E3 is the third member of a production chain that comprises three machines: E1, E2 and E3. Their final goal is to stick LC3 to PE, which will finally make these membranes good for autophagy!
E1, E2, E3 and LC3 work in a timely and highly organised manner: each E1 molecule invests one coin of ATP to load one LC3 tag. As two E1s usually work together, at every cycle two LC3 molecules are loaded. Then, each LC3 is transferred to the cognate E2s, in close proximity to the E1s. In the last step, LC3 is transferred to PE by E3, which works like a tower crane. D. Fracchiolla discovered that in this step WIPI2 plays a fundamental role as it correctly directs and best orients the E3 machine on the membrane making this process very efficient. WIPI2 and E3 can successfully work in a very coordinated manner!
Once the bag has wrapped around the garbage (not described here), the LC3 tag is removed from it. This step is carried by a molecular scissor, called ATG4 which goes all around to trim off LC3 molecules from the bag, so that free LC3 can be re-used in the next bag production cycle!
By joining forces, scientists in the laboratories of Prof. S. Martens and Prof. J. H. Hurley have produced all these molecular machines and have them make work in the test tube. This has allowed them to discover these molecular events that have never been directly observed in living cells, deepening our understanding of life!