Structural Biochemistry/Cell Organelles/Animal Cell/Membrane Contact Site

Overview
Two organelles in a cell come in contact via membrane contact sites (MCS). Membrane contact sites govern the signaling and crossing of ions and lipids from one cellular organelle to the next. Even though research on MCS have been going on for over 50 years now, there is still so much more to uncover regarding their function, structure and regulation. To this date, there are multiple studies done on three main contact sites: the nucleus-vacuole junction, mitochondria-ER, and plasma membrane-ER contact sites. Having membrane- bound organelles is a special characteristic for eukaryotic cells, as it allows for specialized functions in different compartments of the cell. Two organelles exchange information through interorganellar or MCS when they are in close proximity. To visualize their communication, a technique such as electron microscopy was employed. This technique first arose in the 1950s when the close apposition of endoplasmic reticulum and mitochondria was first seen. Ever since then, similar electron microscopy techniques were used to visualize contact sites in cells between the endoplasmic reticulum and: mitochondria, vacuoles, Golgi, chloroplasts and plasma membrane. Since most studies were conducted without molecular markers, they usually refer to proximity of around 30 nanometers between two organelles’ membranes. Unfortunately, there is still no excellent 3D model visualization of MCS. However, in the recent years, there is an increase in the studies of the proteins that reside in the membrane sites and molecular determinants needed to create MCS.

The Nucleus-Vacuole Junction (NVJ)
The membrane contact site between the nucleus and the vacuole is known as the nucleus-vacuole junction (NVJ). The primary study is of the yeast cell Saccharomyces cerevisiae, in which the vacuolar membrane, inner nuclear membrane, and the ER membrane is bridged through NVJ. Nucleus-vacuole junctions are the best studied MCS to this date. NVJ is made of vacuolar protein Vac8 and the ER-localized protein Nvj1. If one of these two proteins is missing from deletion of genes, the MCS will not be made and there would not be contact between two organelles. : There have been two cellular functions suggested for the NVJ. The first one is piecemeal microautophagy of the nucleus. This is a process that marks a part of the yeast nucleus for degradation in the vacuole lumen. Piecemeal microautophagy of the nucleus is stimulated during nitrogen or carbon starvation and requires autophagic parts shared between the electrochemical gradient across the vacuolar membrane, the selective cytoplasm to vacuole, and unselective microautophagy. The second suggested function of NVJ is lipid biosynthesis. The two proteins that participate in lipid biosynthesis, Osh1 and Tsc13, reside in the NVJ. Osh1 comes from the yeast oxysterol-binding protein homology family whose role includes lipid sensors and lipid transfer. However, there is no evidence that shows NVJ regulates lipid biosynthesis because localization of osh1 in NVJ is exclusive to stationary phase. In addition, OSH1 is not needed for forming NVJs or for the piecemeal microautophagy of the nucleus in starved cells. The other protein resides in NVJ (Tsc13) is responsible for catalyzing the terminal step in biosynthesis of very-long-chain fatty acids(VLCFAs). These fatty acids can affect the structure and fluidity of membranes. Although we have pretty good understanding of these two enzymes, we haven't figured out why they reside in NVJ or if there are other proteins that are in the NVJ. : Similar contact sites between ER and the endosome/lysosome (for eukaryote cells) have been observed. Instead of osh1, late endosome-localized oxysterol-binding protein-related protein 1L(ORP1L) works as a cholesterol sensor and it sense level of cholesterol by changing its conformation. Through the change of conformation, ORP1L tells ER how much cholesterol there is in the cell. : Although NVJ is the most studied membrane contact site, we still don't know how much it can do for a cell.

The Mitochondria-ER contact site
The second most-studied MCS is the mitochondria-ER site. This site is primarily visualized via electron microscopy. The section of ER that co-purifies with the mitochondria, known as mitochondria-associated membrane fraction (MAM), is researched extensively on. The two main functions of this site is lipid biosynthesis and Ca2+ homeostasis. Early research showed that the MAM fraction encompasses enzymes that participate in phospholipid biosynthesis. In addition, the mitochondria and ER must work together to make certain lipids. For example, let’s take a closer look in the biosynthetic pathway of phosphatidylcholine. The substrate for this pathway is made in the ER, which then enters the intermembrane space of the mitochondria to produce phosphatidylethanolamine (PE), and then PE goes back to the ER to synthesize phosphatidylcholine. This process shows that the ER and mitochondria collaborate to produce certain lipids. On the other hand, ER-mitochondria MSCs is responsible for the transmission of Ca2+. Homeostasis of mitochondrial Ca2+ is important because Ca2+ regulated the ATP production rate. Furthermore, an overload of Ca2+ can trigger cell death.

The PM-ER contact site
There are many contact sites of ER-PM in different cell types. The function of ER-plasma membrane contact sites is similar to that of the mitochondria-ER junctions, since Ca2+ homeostasis and lipid synthesis and trafficking take place there. As mentioned above, Ca2+ regulation is crucial as it is also responsible for protein synthesis, folding, and signaling. In excitable cells, the global calcium signal is generated by the coupling of PM depolarization and ER calcium release. In non-excitable cells, calcium influx is controlled by detecting luminal ER Ca2+ levels. Research has showed that the PM-ER contact site is involved in non-vesicular lipid trafficking. We know that lipids are insoluble in water. So to transfer lipid from its synthesis sites to its desired work place, one must shield the lipid. In fact, couple families of lipid transfer protein (LTPs) that can perform this task have been found on the contact sites. One of them is oxysterol-binding protein (OSBP) related proteins(ORPS). The oxysterol-binding protein related protein is of a big LTP family which is conserved from yeast to man. It binds to sterols and scientist believe it helps to transfer non-vesicular sterol between the ER and PM. Even though PM-ER contact sites is crucial for Ca2+ and lipid signaling,  we still cannot identify the tether proteins that mediate such contact. One strong possible protein for PM-ER contact sites is a class of highly conserved protein called junctophilins. It can stabilize the junctions by anchoring ER/SR to the PM, giving a structual basis for physiological coupling between PM and ER/SR Ca2+ channels. Further investigation is necessary to map the molecular machinery for the regulation and generation of PM-ER contact sites.

Conclusion of the Membrane Contact Sites (MCS)
Functions of MCS was established first as a site of trafficking of lipids and ions. However, as more and more studies done on MCS, we know that MCSs are also responsible for signaling, effective transmission of metabolic cues within cell, organellar inheritance control mechanism. We are not clear on how MCS perform these tasks yet but better understanding of MCS would be achieved once we figure out its molecular components.