[MUSIC] Hi, Anne-Claude Gavin, I am a biochemist by training and I lead a research group working on lipid metabolism and doing basic research on this topic. We currently know relatively well all the steps and enzymes that are involved in the synthesis of the 2,000 or 3,000 lipid species that are made in human. However, we still understand very poorly how this metabolic network are organized in space in human cells. And this is what keeps our group busy and also what we will be discussing today. So let's zoom in on a membrane bilayer. This is where lipids live, and you see here this yellow lipid. This is where this lipid is made. This is where it will be further metabolized and also where it has very important signaling functions. So in this environment leader relatively free to diffuse laterally membranes are soluble, are flexible and so this lateral diffusion can take place spontaneously. There is no need for a very specialized cellular machinery. However, what lipids cannot do is to freely jump from one side of the leaflet to the other side. And the so called flip flop mechanism require the activity of specialized enzyme that can transfer a lipid from one side to the other side of the membrane. And it's important to realize that without the cellular machinery the lipids would remain stuck at the places at the site where they have been synthesized. For example, in the plasma membrane we know that lipid accumulate in the inner leaflet. For example, phosphatidylserine or phosphatidylethanolamine. And this is the result of the activity of enzyme cause called flipases. So they create asymmetry across the membrane bilayer. And this is key for self functioning and cell biology. In contrast, the ER has a very rapid distribution of most lipid species across the two bilayer. This is the activity of scramblases. And here the scramblases scramble lipids, to open the gate and alow a very rapid distribution across the bilayer. In some cases those events require energy, require hydrolysis of ATP. This is especially true for the creation of asymmetry across the bi-bilayer. When lipid needs to be bumped against a concentration gradient, but has, we've seen this really vary as a function of the organelle. Another thing that lipids cannot do easily, it is to jump out of the membrane bilayer. Lipid are hydrophobic so they cannot freely diffuse in the cytoplasm. So again, this means that a lipid made in the membrane of a specific organelle is not free to jump to the membrane of a another organelle. So without a specialized cellular machinery, lipids would remain blocked at the place where they have been made. So let's see where are lipids are made. So the main places where most lipid synthesis starts is the endoplasmic reticulum (ER). But then later on those lipid species are required in all the different organelles or the different membranes. Either for further metabolism, for signaling function or to define the identity of those organelles. So a good example is the phosphatidylinositol. It is made in the endoplasmic reticulum, but then this lipid is required in many other places. It gets phosphorylated in the different organelle by the activity of specific kinases that decorate the inositol group with phosphate at specific position(s). This is, for example, the case in the Golgi, where domain species of phosphatidylinositol-4-phosphate. But we also have the plasma membrane species and the endosomal species. And they define the identity of the different organellar membranes. So the system requires a lipid transport. Another example is phosphatidylserine. Again it is made in the endoplasmic reticulum, but then this lipid is required in the plasma membrane, where it accumulates in the inner leaflet. And it has very important signaling functions. So we here the transport of phosphatidylserine to the plasma membrane. Can import take place via vesicular trafficking via exocytosis. However, phosphatidylserine is also required in the mitochondria. Where it is further required for metabolism, for decarboxylation to another, lipid phosphatidylethanolamine. However, the mitochondria is not part of the vesicular trafficking machinery, so this means that additional mechanisms are required that fulfilled this transfer function. So this is really something critical for the cell. We have seen that lipids are hydrophobic, they cannot freely diffuse out of the membranes. We have seen that they're very often synthesized at places that are very far away from the places where they have important biological functions, and this is signaling and metabolism. So the cell needs to have a logistical system that can dispatch lipid at the proper place and at the proper time. So we know that families of protein, lipid transfer protein have evolved that can fulfill some of those functions. So these are very sophisticated biochemical small machines that can sense at donor Membrane. This is critical, they want to take the lipid from the proper place in this membrane. They can extract their lipid cargo and then they must lose affinity for the donor membrane on gain affinity for the acceptor membrane. Again, this is critical, you don't want to deliver the lipid randomly or at the wrong place. Then the lipid is delivered to this acceptor membrane. The protein loses its affinity for the acceptor membrane and gain back affinity for the donor membrane and cycle like that. So we know more than 130 differently lipid transfer protein in human. They can be classified into ten families according to the structure of their lipid transfer domain. And in human anything that will interfere with the function of those protein like mutation will lead to disorder. An example is the Niemann-Pick syndrome, where we see the accumulation of cholesterol in an organelle where usually cholesterol does not accumulate, and this is lysosome. So these proteins have a very different lipid transfer domain, but they have a very common mode of action and this is that they can encapsulate and hydrophobic ligands. Here a fatty acid in and hydrophobic pocket, and you can see the hydrophobic residue that flanked the binding side tryptophan and tyrosine's. And the outside of the protein is hydrophilic, so that it is soluble in the cytoplasm and can bring the lipid in the aqueous phases of the cell. So this is a very common mode of action. They have an hydrophobic center and a hydrophilic exterior. And this is very important for their mode of action, they need to bind lipids very tightly, but they also need to be able to deliver the lipids so these binding activity must be strong but reversible. In some cases we also see that this protein also have a lid that can lock the lipid in its binding pocket. In this example, you see a phosphatidylserine and you see the head group colored in red that makes contact with a lid in green. Unlock the phosphatidylserine inside the binding pocket. And this mechanism is key for the controlled release of the lipid in the proper membrane in the proper environment. In the majority of the cases [those] lipid transfer proteins bind lipids one at a time. But you see here a beautiful exception and this is an example where the lipid transfer protein binds both cholesterol ester and triacylglycerols. So almost all lipids species are carried by lipid transfer proteins. Interestingly, the same lipid can be transported by different carriers and reciprocally, the same carrier can also transport different lipid species. And what I found very, very inspiring is the fact that in the majority of the cases we still don't understand what cargo those transporters transport. So this means that there is still a lot of research to be done, and that keeps us very busy. So a few words about the localization of those transporters in the cell. You see the LTP here, the 130 classified. According to their belonging to families and you can see that the main organelle contain more than one lipid transporter. So the decorate virtually all organelles inside the cell. And they localize in very specialized membrane area that are called membrane contact sites. They link the endoplasmic reticulum, the lipid factory of the cell, to virtually all other membranes in the cell. And they come in very close proximity, about 20 nanometer. So many different type of membrane contact site, but here we'll focus on the one between the On the mitochondria. And you see here electron micrograph of those contact sites. You see dark dots that are the ribosomes and they decorate an organelle. This is the endo plasmic reticulum so the rough endoplasmic reticulum. They also get lost at some point on this becomes the smooth endoplasmic reticulum. So we really see different type of membrane here, the endoplasmic reticulum and the mitochondria. And in the zoom in image on the right, you see that those membrane come in very close proximity, about 20 nanometer. It makes like a zipper and these places are the membrane contact sites. This is where lipid metabolism takes place and also lipid transport. So to summarize what we have learned today, we have seen that lipids are very frequently made in places that are very remote from the places where those lipids play important biological function. This means metabolism and also signaling. And that lipid are insoluble, so they're not free to diffuse and jump from different membrane. So systems are needed inside the cell to transport the lipid at the proper place, but also at the proper time. And that specialized proteins, called lipid transfer proteins, can assist with those functions. And we've seen that this protein localize in very specialized areas that are called membrane contact sites. [MUSIC]
