Lipid bilayer

The lipid bilayer (or phospholipid bilayer) is a thin polar membrane made of two layers of lipid molecules. These membranes are flat sheets that form a continuous barrier around all cells. The cell membranes of almost all organisms and many viruses are made of a lipid bilayer, as are the nuclear membrane surrounding the cell nucleus, and membranes of the membrane-bound organelles in the cell. The lipid bilayer is the barrier that keeps ions, proteins and other molecules where they are needed and prevents them from diffusing into areas where they should not be. Lipid bilayers are ideally suited to this role, even though they are only a few nanometers in width, because they are impermeable to most water-soluble (hydrophilic) molecules. Bilayers are particularly impermeable to ions, which allows cells to regulate salt concentrations and pH by transporting ions across their membranes using proteins called ion pumps.
Biological bilayers are usually composed of amphiphilic phospholipids that have a hydrophilic phosphate head and a hydrophobic tail consisting of two fatty acid chains. Phospholipids with certain head groups can alter the surface chemistry of a bilayer and can, for example, serve as signals as well as "anchors" for other molecules in the membranes of cells. Just like the heads, the tails of lipids can also affect membrane properties, for instance by determining the phase of the bilayer. The bilayer can adopt a solid gel phase state at lower temperatures but undergo phase transition to a fluid state at higher temperatures, and the chemical properties of the lipids' tails influence at which temperature this happens. The packing of lipids within the bilayer also affects its mechanical properties, including its resistance to stretching and bending. Many of these properties have been studied with the use of artificial "model" bilayers produced in a lab. Vesicles made by model bilayers have also been used clinically to deliver drugs.
Biological membranes typically include several types of molecules other than phospholipids. A particularly important example in animal cells is cholesterol, which helps strengthen the bilayer and decrease its permeability. Cholesterol also helps regulate the activity of certain integral membrane proteins. Integral membrane proteins function when incorporated into a lipid bilayer, and they are held tightly to the lipid bilayer with the help of an annular lipid shell. Because bilayers define the boundaries of the cell and its compartments, these membrane proteins are involved in many intra- and inter-cellular signaling processes. Certain kinds of membrane proteins are involved in the process of fusing two bilayers together. This fusion allows the joining of two distinct structures as in the acrosome reaction during fertilization of an egg by a sperm, or the entry of a virus into a cell. Because lipid bilayers are fragile and invisible in a traditional microscope, they are a challenge to study. Experiments on bilayers often require advanced techniques like electron microscopy and atomic force microscopy.
Structure and organization.
When phospholipids are exposed to water, they self-assemble into a two-layered sheet with the hydrophobic tails pointing toward the center of the sheet. This arrangement results in two “leaflets” that are each a single molecular layer. The center of this bilayer contains almost no water and excludes molecules like sugars or salts that dissolve in water. The assembly process is driven by interactions between hydrophobic molecules (also called the hydrophobic effect). An increase in interactions between hydrophobic molecules (causing clustering of hydrophobic regions) allows water molecules to bond more freely with each other, increasing the entropy of the system. This complex process includes non-covalent interactions such as van der Waals forces, electrostatic and hydrogen bonds.
Cross section analysis.
The lipid bilayer is very thin compared to its lateral dimensions. If a typical mammalian cell (diameter ~10 micrometers) were magnified to the size of a watermelon (~1 ft/30 cm), the lipid bilayer making up the plasma membrane would be about as thick as a piece of office paper. Despite being only a few nanometers thick, the bilayer is composed of several distinct chemical regions across its cross-section. These regions and their interactions with the surrounding water have been characterized over the past several decades with x-ray reflectometry, neutron scattering and nuclear magnetic resonance techniques.
The first region on either side of the bilayer is the hydrophilic headgroup. This portion of the membrane is completely hydrated and is typically around 0.8-0.9 nm thick. In phospholipid bilayers the phosphate group is located within this hydrated region, approximately 0.5 nm outside the hydrophobic core. In some cases, the hydrated region can extend much further, for instance in lipids with a large protein or long sugar chain grafted to the head. One common example of such a modification in nature is the lipopolysaccharide coat on a bacterial outer membrane, which helps retain a water layer around the bacterium to prevent dehydration.
Next to the hydrated region is an intermediate region that is only partially hydrated. This boundary layer is approximately 0.3 nm thick. Within this short distance, the water concentration drops from 2M on the headgroup side to nearly zero on the tail (core) side. The hydrophobic core of the bilayer is typically 3-4 nm thick, but this value varies with chain length and chemistry. Core thickness also varies significantly with temperature, in particular near a phase transition.
Asymmetry.
In many naturally occurring bilayers, the compositions of the inner and outer membrane leaflets are different. In human red blood cells, the inner (cytoplasmic) leaflet is composed mostly of phosphatidylethanolamine, phosphatidylserine and phosphatidylinositol and its phosphorylated derivatives. By contrast, the outer (extracellular) leaflet is based on phosphatidylcholine, sphingomyelin and a variety of glycolipids. In some cases, this asymmetry is based on where the lipids are made in the cell and reflects their initial orientation. The biological functions of lipid asymmetry are imperfectly understood, although it is clear that it is used in several different situations. For example, when a cell undergoes apoptosis, the phosphatidylserine — normally localised to the cytoplasmic leaflet — is transferred to the outer surface: There, it is recognised by a macrophage that then actively scavenges the dying cell.
Lipid asymmetry arises, at least in part, from the fact that most phospholipids are synthesised and initially inserted into the inner monolayer: those that constitute the outer monolayer are then transported from the inner monolayer by a class of enzymes called flippases. Other lipids, such as sphingomyelin, appear to be synthesised at the external leaflet. Flippases are members of a larger family of lipid transport molecules that also includes floppases, which transfer lipids in the opposite direction, and scramblases, which randomize lipid distribution across lipid bilayers (as in apoptotic cells). In any case, once lipid asymmetry is established, it does not normally dissipate quickly because spontaneous flip-flop of lipids between leaflets is extremely slow.
It is possible to mimic this asymmetry in the laboratory in model bilayer systems. Certain types of very small artificial vesicle will automatically make themselves slightly asymmetric, although the mechanism by which this asymmetry is generated is very different from that in cells. By utilizing two different monolayers in Langmuir-Blodgett deposition or a combination of Langmuir-Blodgett and vesicle rupture deposition it is also possible to synthesize an asymmetric planar bilayer. This asymmetry may be lost over time as lipids in supported bilayers can be prone to flip-flop.
Phases and phase transitions.
At a given temperature a lipid bilayer can exist in either a liquid or a gel (solid) phase. All lipids have a characteristic temperature at which they transition (melt) from the gel to liquid phase. In both phases the lipid molecules are prevented from flip-flopping across the bilayer, but in liquid phase bilayers a given lipid will exchange locations with its neighbor millions of times a second. This random walk exchange allows lipid to diffuse and thus wander across the surface of the membrane. Unlike liquid phase bilayers, the lipids in a gel phase bilayer have less mobility.
The phase behavior of lipid bilayers is determined largely by the strength of the attractive Van der Waals interactions between adjacent lipid molecules. Longer-tailed lipids have more area over which to interact, increasing the strength of this interaction and, as a consequence, decreasing the lipid mobility. Thus, at a given temperature, a short-tailed lipid will be more fluid than an otherwise identical long-tailed lipid. Transition temperature can also be affected by the degree of unsaturation of the lipid tails. An unsaturated double bond can produce a kink in the alkane chain, disrupting the lipid packing. This disruption creates extra free space within the bilayer that allows additional flexibility in the adjacent chains. An example of this effect can be noted in everyday life as butter, which has a large percentage saturated fats, is solid at room temperature while vegetable oil, which is mostly unsaturated, is liquid.
Most natural membranes are a complex mixture of different lipid molecules. If some of the components are liquid at a given temperature while others are in the gel phase, the two phases can coexist in spatially separated regions, rather like an iceberg floating in the ocean. This phase separation plays a critical role in biochemical phenomena because membrane components such as proteins can partition into one or the other phase and thus be locally concentrated or activated. One particularly important component of many mixed phase systems is cholesterol, which modulates bilayer permeability, mechanical strength, and biochemical interactions.
Surface chemistry.
While lipid tails primarily modulate bilayer phase behavior, it is the headgroup that determines the bilayer surface chemistry. Most natural bilayers are composed primarily of phospholipids, but sphingolipids and sterols such as cholesterol are also important components. Of the phospholipids, the most common headgroup is phosphatidylcholine (PC), accounting for about half the phospholipids in most mammalian cells. PC is a zwitterionic headgroup, as it has a negative charge on the phosphate group and a positive charge on the amine but, because these local charges balance, no net charge.
Other headgroups are also present to varying degrees and can include phosphatidylserine (PS) phosphatidylethanolamine (PE) and phosphatidylglycerol (PG). These alternate headgroups often confer specific biological functionality that is highly context-dependent. For instance, PS presence on the extracellular membrane face of erythrocytes is a marker of cell apoptosis, whereas PS in growth plate vesicles is necessary for the nucleation of hydroxyapatite crystals and subsequent bone mineralization. Unlike PC, some of the other headgroups carry a net charge, which can alter the electrostatic interactions of small molecules with the bilayer.
Biological roles.
Containment and separation.
The primary role of the lipid bilayer in biology is to separate aqueous compartments from their surroundings. Without some form of barrier delineating “self” from “non-self”, it is difficult to even define the concept of an organism or of life. This barrier takes the form of a lipid bilayer in all known life forms except for a few species of archaea that utilize a specially adapted lipid monolayer. It has even been proposed that the very first form of life may have been a simple lipid vesicle with virtually its sole biosynthetic capability being the production of more phospholipids. The partitioning ability of the lipid bilayer is based on the fact that hydrophilic molecules cannot easily cross the hydrophobic bilayer core, as discussed in Transport across the bilayer below. The nucleus, mitochondria and chloroplasts have two lipid bilayers, while other sub-cellular structures are surrounded by a single lipid bilayer (such as the plasma membrane, endoplasmic reticula, Golgi apparatus and lysosomes). See Organelle.
Prokaryotes have only one lipid bilayer - the cell membrane (also known as the plasma membrane). Many prokaryotes also have a cell wall, but the cell wall is composed of proteins or long chain carbohydrates, not lipids. In contrast, eukaryotes have a range of organelles including the nucleus, mitochondria, lysosomes and endoplasmic reticulum. All of these sub-cellular compartments are surrounded by one or more lipid bilayers and, together, typically comprise the majority of the bilayer area present in the cell. In liver hepatocytes for example, the plasma membrane accounts for only two percent of the total bilayer area of the cell, whereas the endoplasmic reticulum contains more than fifty percent and the mitochondria a further thirty percent.
Signaling.
Probably the most familiar form of cellular signaling is synaptic transmission, whereby a nerve impulse that has reached the end of one neuron is conveyed to an adjacent neuron via the release of neurotransmitters. This transmission is made possible by the action of synaptic vesicles loaded with the neurotransmitters to be released. These vesicles fuse with the cell membrane at the pre-synaptic terminal and release its contents to the exterior of the cell. The contents then diffuse across the synapse to the post-synaptic terminal.
Lipid bilayers are also involved in signal transduction through their role as the home of integral membrane proteins. This is an extremely broad and important class of biomolecule. It is estimated that up to a third of the human proteome are membrane proteins. Some of these proteins are linked to the exterior of the cell membrane. An example of this is the CD59 protein, which identifies cells as “self” and thus inhibits their destruction by the immune system. The HIV virus evades the immune system in part by grafting these proteins from the host membrane onto its own surface. Alternatively, some membrane proteins penetrate all the way through the bilayer and serve to relay individual signal events from the outside to the inside of the cell. The most common class of this type of protein is the G protein-coupled receptor (GPCR). GPCRs are responsible for much of the cell's ability to sense its surroundings and, because of this important role, approximately 40% of all modern drugs are targeted at GPCRs.
In addition to protein- and solution-mediated processes, it is also possible for lipid bilayers to participate directly in signaling. A classic example of this is phosphatidylserine-triggered phagocytosis. Normally, phosphatidylserine is asymmetrically distributed in the cell membrane and is present only on the interior side. During programmed cell death a protein called a scramblase equilibrates this distribution, displaying phosphatidylserine on the extracellular bilayer face. The presence of phosphatidylserine then triggers phagocytosis to remove the dead or dying cell.
Characterization methods.
The lipid bilayer is a very difficult structure to study because it is so thin and fragile. In spite of these limitations dozens of techniques have been developed over the last seventy years to allow investigations of its structure and function.
Electrical measurements.
Electrical measurements are a straightforward way to characterize an important function of a bilayer: its ability to segregate and prevent the flow of ions in solution. By applying a voltage across the bilayer and measuring the resulting current, the resistance of the bilayer is determined. This resistance is typically quite high (108 Ohm-cm2 or more) since the hydrophobic core is impermeable to charged species. The presence of even a few nanometer-scale holes results in a dramatic increase in current. The sensitivity of this system is such that even the activity of single ion channels can be resolved.
Fluorescence microscopy.
Electrical measurements do not provide an actual picture like imaging with a microscope can. Lipid bilayers cannot be seen in a traditional microscope because they are too thin. In order to see bilayers, researchers often use fluorescence microscopy. A sample is excited with one wavelength of light and observed in a different wavelength, so that only fluorescent molecules with a matching excitation and emission profile will be seen. Natural lipid bilayers are not fluorescent, so a dye is used that attaches to the desired molecules in the bilayer. Resolution is usually limited to a few hundred nanometers, much smaller than a typical cell but much larger than the thickness of a lipid bilayer.
Electron microscopy.
Electron microscopy offers a higher resolution image. In an electron microscope, a beam of focused electrons interacts with the sample rather than a beam of light as in traditional microscopy. In conjunction with rapid freezing techniques, electron microscopy has also been used to study the mechanisms of inter- and intracellular transport, for instance in demonstrating that exocytotic vesicles are the means of chemical release at synapses.
Nuclear magnetic resonance spectroscopy.
31P-NMR(nuclear magnetic resonance) spectroscopy is widely used for studies of phospholipid bilayers and biological membranes in native conditions. The analysis of 31P-NMR spectra of lipids could provide a wide range of information about lipid bilayer packing, phase transitions (gel phase, physiological liquid crystal phase, ripple phases, non bilayer phases), lipid head group orientation/dynamics, and elastic properties of pure lipid bilayer and as a result of binding of proteins and other biomolecules.
Atomic force microscopy.
A new method to study lipid bilayers is Atomic force microscopy (AFM). Rather than using a beam of light or particles, a very small sharpened tip scans the surface by making physical contact with the bilayer and moving across it, like a record player needle. AFM is a promising technique because it has the potential to image with nanometer resolution at room temperature and even under water or physiological buffer, conditions necessary for natural bilayer behavior. Utilizing this capability, AFM has been used to examine dynamic bilayer behavior including the formation of transmembrane pores (holes) and phase transitions in supported bilayers. Another advantage is that AFM does not require fluorescent or isotopic labeling of the lipids, since the probe tip interacts mechanically with the bilayer surface. Because of this, the same scan can image both lipids and associated proteins, sometimes even with single-molecule resolution. AFM can also probe the mechanical nature of lipid bilayers.
Dual polarisation interferometry.
Lipid bilayers exhibit high levels of birefringence where the refractive index in the plane of the bilayer differs from that perpendicular by as much as 0.1 refractive index units. This has been used to characterise the degree of order and disruption in bilayers using dual polarisation interferometry to understand mechanisms of protein interaction.
Quantum chemical calculations.
Lipid bilayers are complicated molecular systems with many degrees of freedom. Thus, atomistic simulation of membrane and in particular ab initio calculations of its properties is difficult and computationally expensive. Quantum chemical calculations has recently been successfully performed to estimate dipole and quadrupole moments of lipid membranes.
Transport across the bilayer.
Passive diffusion.
Most polar molecules have low solubility in the hydrocarbon core of a lipid bilayer and, as a consequence, have low permeability coefficients across the bilayer. This effect is particularly pronounced for charged species, which have even lower permeability coefficients than neutral polar molecules. Anions typically have a higher rate of diffusion through bilayers than cations. Compared to ions, water molecules actually have a relatively large permeability through the bilayer, as evidenced by osmotic swelling. When a cell or vesicle with a high interior salt concentration is placed in a solution with a low salt concentration it will swell and eventually burst. Such a result would not be observed unless water was able to pass through the bilayer with relative ease. The anomalously large permeability of water through bilayers is still not completely understood and continues to be the subject of active debate. Small uncharged apolar molecules diffuse through lipid bilayers many orders of magnitude faster than ions or water. This applies both to fats and organic solvents like chloroform and ether. Regardless of their polar character larger molecules diffuse more slowly across lipid bilayers than small molecules.
Ion pumps and channels.
Two special classes of protein deal with the ionic gradients found across cellular and sub-cellular membranes in nature- ion channels and ion pumps. Both pumps and channels are integral membrane proteins that pass through the bilayer, but their roles are quite different. Ion pumps are the proteins that build and maintain the chemical gradients by utilizing an external energy source to move ions against the concentration gradient to an area of higher chemical potential. The energy source can be ATP, as is the case for the Na+-K+ ATPase. Alternatively, the energy source can be another chemical gradient already in place, as in the Ca2+/Na+ antiporter. It is through the action of ion pumps that cells are able to regulate pH via the pumping of protons.
In contrast to ion pumps, ion channels do not build chemical gradients but rather dissipate them in order to perform work or send a signal. Probably the most familiar and best studied example is the voltage-gated Na+ channel, which allows conduction of an action potential along neurons. All ion pumps have some sort of trigger or “gating” mechanism. In the previous example it was electrical bias, but other channels can be activated by binding a molecular agonist or through a conformational change in another nearby protein.
Endocytosis and exocytosis.
Some molecules or particles are too large or too hydrophilic to pass through a lipid bilayer. Other molecules could pass through the bilayer but must be transported rapidly in such large numbers that channel-type transport is impractical. In both cases, these types of cargo can be moved across the cell membrane through fusion or budding of vesicles. When a vesicle is produced inside the cell and fuses with the plasma membrane to release its contents into the extracellular space, this process is known as exocytosis. In the reverse process, a region of the cell membrane will dimple inwards and eventually pinch off, enclosing a portion of the extracellular fluid to transport it into the cell. Endocytosis and exocytosis rely on very different molecular machinery to function, but the two processes are intimately linked and could not work without each other. The primary mechanism of this interdependence is the large amount of lipid material involved. In a typical cell, an area of bilayer equivalent to the entire plasma membrane will travel through the endocytosis/exocytosis cycle in about half an hour. If these two processes were not balancing each other, the cell would either balloon outward to an unmanageable size or completely deplete its plasma membrane within a short time.
Exocytosis in prokaryotes: Membrane vesicular exocytosis, popularly known as membrane vesicle trafficking, a Nobel prize-winning (year, 2013) process, is traditionally regarded as a prerogative of eukaryotic cells. This "myth" was however broken with the revelation that nanovesicles, popularly known as bacterial outer membrane vesicles, released by gram-negative microbes, translocate bacterial signal molecules to host or target cells to carry out multiple processes in favour of the secreting microbe e.g., in "host cell invasion" and microbe-environment interactions, in general.
Electroporation.
Electroporation is the rapid increase in bilayer permeability induced by the application of a large artificial electric field across the membrane. Experimentally, electroporation is used to introduce hydrophilic molecules into cells. It is a particularly useful technique for large highly charged molecules such as DNA, which would never passively diffuse across the hydrophobic bilayer core. Because of this, electroporation is one of the key methods of transfection as well as bacterial transformation. It has even been proposed that electroporation resulting from lightning strikes could be a mechanism of natural horizontal gene transfer.
This increase in permeability primarily affects transport of ions and other hydrated species, indicating that the mechanism is the creation of nm-scale water-filled holes in the membrane. Although electroporation and dielectric breakdown both result from application of an electric field, the mechanisms involved are fundamentally different. In dielectric breakdown the barrier material is ionized, creating a conductive pathway. The material alteration is thus chemical in nature. In contrast, during electroporation the lipid molecules are not chemically altered but simply shift position, opening up a pore that acts as the conductive pathway through the bilayer as it is filled with water.
Mechanics.
Lipid bilayers are large enough structures to have some of the mechanical properties of liquids or solids. The area compression modulus Ka, bending modulus Kb, and edge energy formula_1, can be used to describe them. Solid lipid bilayers also have a shear modulus, but like any liquid, the shear modulus is zero for fluid bilayers. These mechanical properties affect how the membrane functions. Ka and Kb affect the ability of proteins and small molecules to insert into the bilayer, and bilayer mechanical properties have been shown to alter the function of mechanically activated ion channels. Bilayer mechanical properties also govern what types of stress a cell can withstand without tearing. Although lipid bilayers can easily bend, most cannot stretch more than a few percent before rupturing.
As discussed in the Structure and organization section, the hydrophobic attraction of lipid tails in water is the primary force holding lipid bilayers together. Thus, the elastic modulus of the bilayer is primarily determined by how much extra area is exposed to water when the lipid molecules are stretched apart. It is not surprising given this understanding of the forces involved that studies have shown that Ka varies strongly with osmotic pressure but only weakly with tail length and unsaturation. Because the forces involved are so small, it is difficult to experimentally determine Ka. Most techniques require sophisticated microscopy and very sensitive measurement equipment.
In contrast to Ka, which is a measure of how much energy is needed to stretch the bilayer, Kb is a measure of how much energy is needed to bend or flex the bilayer. Formally, bending modulus is defined as the energy required to deform a membrane from its intrinsic curvature to some other curvature. Intrinsic curvature is defined by the ratio of the diameter of the head group to that of the tail group. For two-tailed PC lipids, this ratio is nearly one so the intrinsic curvature is nearly zero. If a particular lipid has too large a deviation from zero intrinsic curvature it will not form a bilayer and will instead form other phases such as micelles or inverted micelles. Addition of "small hydrophilic molecules" like "sucrose" into mixed lipid "lamellar liposomes" made from galactolipid-rich thylakoid membranes destabilises bilayers into micellar phase. Typically, Kb is not measured experimentally but rather is calculated from measurements of Ka and bilayer thickness, since the three parameters are related.
formula_1 is a measure of how much energy it takes to expose a bilayer edge to water by tearing the bilayer or creating a hole in it. The origin of this energy is the fact that creating such an interface exposes some of the lipid tails to water, but the exact orientation of these border lipids is unknown. There is some evidence that both hydrophobic (tails straight) and hydrophilic (heads curved around) pores can coexist.
Fusion.
Fusion is the process by which two lipid bilayers merge, resulting in one connected structure. If this fusion proceeds completely through both leaflets of both bilayers, a water-filled bridge is formed and the solutions contained by the bilayers can mix. Alternatively, if only one leaflet from each bilayer is involved in the fusion process, the bilayers are said to be hemifused. Fusion is involved in many cellular processes, in particular in eukaryotes, since the eukaryotic cell is extensively sub-divided by lipid bilayer membranes. Exocytosis, fertilization of an egg by sperm activation, and transport of waste products to the lysozome are a few of the many eukaryotic processes that rely on some form of fusion. Even the entry of pathogens can be governed by fusion, as many bilayer-coated viruses have dedicated fusion proteins to gain entry into the host cell.
There are four fundamental steps in the fusion process. First, the involved membranes must aggregate, approaching each other to within several nanometers. Second, the two bilayers must come into very close contact (within a few angstroms). To achieve this close contact, the two surfaces must become at least partially dehydrated, as the bound surface water normally present causes bilayers to strongly repel. The presence of ions, in particular divalent cations like magnesium and calcium, strongly affects this step. One of the critical roles of calcium in the body is regulating membrane fusion. Third, a destabilization must form at one point between the two bilayers, locally distorting their structures. The exact nature of this distortion is not known. One theory is that a highly curved "stalk" must form between the two bilayers. Proponents of this theory believe that it explains why phosphatidylethanolamine, a highly curved lipid, promotes fusion. Finally, in the last step of fusion, this point defect grows and the components of the two bilayers mix and diffuse away from the site of contact.
The situation is further complicated when considering fusion "in vivo" since biological fusion is almost always regulated by the action of membrane-associated proteins. The first of these proteins to be studied were the viral fusion proteins, which allow an enveloped virus to insert its genetic material into the host cell (enveloped viruses are those surrounded by a lipid bilayer; some others have only a protein coat). Eukaryotic cells also use fusion proteins, the best-studied of which are the SNAREs. SNARE proteins are used to direct all vesicular intracellular trafficking. Despite years of study, much is still unknown about the function of this protein class. In fact, there is still an active debate regarding whether SNAREs are linked to early docking or participate later in the fusion process by facilitating hemifusion.
In studies of molecular and cellular biology it is often desirable to artificially induce fusion. The addition of polyethylene glycol (PEG) causes fusion without significant aggregation or biochemical disruption. This procedure is now used extensively, for example by fusing B-cells with myeloma cells. The resulting “hybridoma” from this combination expresses a desired antibody as determined by the B-cell involved, but is immortalized due to the melanoma component. Fusion can also be artificially induced through electroporation in a process known as electrofusion. It is believed that this phenomenon results from the energetically active edges formed during electroporation, which can act as the local defect point to nucleate stalk growth between two bilayers.
Model systems.
Lipid bilayers can be created artificially in the lab to allow researchers to perform experiments that cannot be done with natural bilayers. They can also be used in the field of Synthetic Biology, to define the boundaries of artificial cells. These synthetic systems are called model lipid bilayers. There are many different types of model bilayers, each having experimental advantages and disadvantages. They can be made with either synthetic or natural lipids. Among the most common model systems are:
Commercial applications.
To date, the most successful commercial application of lipid bilayers has been the use of liposomes for drug delivery, especially for cancer treatment. (Note- the term “liposome” is in essence synonymous with “vesicle” except that vesicle is a general term for the structure whereas liposome refers to only artificial not natural vesicles) The basic idea of liposomal drug delivery is that the drug is encapsulated in solution inside the liposome then injected into the patient. These drug-loaded liposomes travel through the system until they bind at the target site and rupture, releasing the drug. In theory, liposomes should make an ideal drug delivery system since they can isolate nearly any hydrophilic drug, can be grafted with molecules to target specific tissues and can be relatively non-toxic since the body possesses biochemical pathways for degrading lipids.
The first generation of drug delivery liposomes had a simple lipid composition and suffered from several limitations. Circulation in the bloodstream was extremely limited due to both renal clearing and phagocytosis. Refinement of the lipid composition to tune fluidity, surface charge density, and surface hydration resulted in vesicles that adsorb fewer proteins from serum and thus are less readily recognized by the immune system. The most significant advance in this area was the grafting of polyethylene glycol (PEG) onto the liposome surface to produce “stealth” vesicles, which circulate over long times without immune or renal clearing.
The first stealth liposomes were passively targeted at tumor tissues. Because tumors induce rapid and uncontrolled angiogenesis they are especially “leaky” and allow liposomes to exit the bloodstream at a much higher rate than normal tissue would. More recently work has been undertaken to graft antibodies or other molecular markers onto the liposome surface in the hope of actively binding them to a specific cell or tissue type. Some examples of this approach are already in clinical trials.
Another potential application of lipid bilayers is the field of biosensors. Since the lipid bilayer is the barrier between the interior and exterior of the cell, it is also the site of extensive signal transduction. Researchers over the years have tried to harness this potential to develop a bilayer-based device for clinical diagnosis or bioterrorism detection. Progress has been slow in this area and, although a few companies have developed automated lipid-based detection systems, they are still targeted at the research community. These include Biacore (now GE Healthcare Life Sciences), which offers a disposable chip for utilizing lipid bilayers in studies of binding kinetics and Nanion Inc., which has developed an automated patch clamping system. Other, more exotic applications are also being pursued such as the use of lipid bilayer membrane pores for DNA sequencing by Oxford Nanolabs. To date, this technology has not proven commercially viable.
A supported lipid bilayer (SLB) as described above has achieved commercial success as a screening technique to measure the permeability of drugs. This parallel artificial membrane permeability assay PAMPA technique measures the permeability across specifically formulated lipid cocktail(s) found to be highly correlated with Caco-2 cultures, the gastrointestinal tract, blood–brain barrier and skin.
History.
By the early twentieth century scientists had come to believe that cells are surrounded by a thin oil-like barrier, but the structural nature of this membrane was not known. Two experiments in 1925 laid the groundwork to fill in this gap. By measuring the capacitance of erythrocyte solutions, Hugo Fricke determined that the cell membrane was 3.3 nm thick.
Although the results of this experiment were accurate, Fricke misinterpreted the data to mean that the cell membrane is a single molecular layer. Prof. Dr. Evert Gorter (1881–1954) and F. Grendel of Leiden University approached the problem from a different perspective, spreading the erythrocyte lipids as a monolayer on a Langmuir-Blodgett trough. When they compared the area of the monolayer to the surface area of the cells, they found a ratio of two to one. Later analyses showed several errors and incorrect assumptions with this experiment but, serendipitously, these errors canceled out and from this flawed data Gorter and Grendel drew the correct conclusion- that the cell membrane is a lipid bilayer.
This theory was confirmed through the use of electron microscopy in the late 1950s. Although he did not publish the first electron microscopy study of lipid bilayers J. David Robertson was the first to assert that the two dark electron-dense bands were the headgroups and associated proteins of two apposed lipid monolayers. In this body of work, Robertson put forward the concept of the “unit membrane.” This was the first time the bilayer structure had been universally assigned to all cell membranes as well as organelle membranes.
Around the same time, the development of model membranes confirmed that the lipid bilayer is a stable structure that can exist independent of proteins. By “painting” a solution of lipid in organic solvent across an aperture, Mueller and Rudin were able to create an artificial bilayer and determine that this exhibited lateral fluidity, high electrical resistance and self-healing in response to puncture, all of which are properties of a natural cell membrane. A few years later, Alec Bangham showed that bilayers, in the form of lipid vesicles, could also be formed simply by exposing a dried lipid sample to water. This was an important advance, since it demonstrated that lipid bilayers form spontaneously via self assembly and do not require a patterned support structure.
In 1977, a totally synthetic bilayer membrane was prepared by Kunitake and Okahata, from a single organic compound, didodecyldimethylammonium bromide. It clearly shows that the bilayer membrane was assembled by the van der Waals interaction.


Melanin-concentrating hormone

Melanin-concentrating hormone (MCH) is a cyclic 19-amino acid orexigenic hypothalamic peptide originally isolated from the pituitary gland of teleost fish, where it controls skin pigmentation. In mammals it is involved in the regulation of feeding behavior, mood, sleep-wake cycle and energy balance.
Structure.
MCH is a cyclic 19-amino acid neuropeptide, as it is a polypeptide chain that is able to act as a neurotransmitter. MCH neurons are mainly concentrated in the lateral hypothalamic area, zona incerta, and the incerto-hypothalamic area, but they are also located, in much smaller amounts, in the paramedian pontine reticular formation (PPRF), medial preoptic area, laterodorsal tegmental nucleus, and the olfactory tubercle. MCH is activated by binding to two G-coupled protein receptors (GCPRs), MCHR1 and MCHR2. MCHR2 has only been identified in certain species such as humans, dogs, ferrets, and rhesus monkeys, while other mammals such as rodents and rabbits do not have the receptor. MCH is cleaved from prepro-MCH (ppMCH), a 165 amino acid polypeptide which also contains the neuropeptides GE and EI.
Tissue distribution.
MCH has also been found in peripheral structures outside of the brain. Both the spleen and thymus have shown significant levels of MCH in mammals in multiple studies. The bloodstream seems to carry MCH around the body in mammals as well, though it is a very amount in humans.
MCH is found in the laterodorsal tegmental nucleus solely in female brains in rat models. MCH has also only been found in the medial preoptic area and the paraventricular hypothalamic nucleus during lactation.
Activation and deactivation.
MCH neurons depolarize in response to high glucose concentrations. This mechanism seems to be related to glucose being used as a reactant to form ATP, which also causes MCH neurons to depolarize. The neurotransmitter, glutamate, also causes MCH neurons to depolarize, while another neurotransmitter, GABA, causes MCH neurons to hyperpolarize. Orexin also depolarizes MCH neurons. MCH neurons seems to have an inhibitory response to MCH, but does not cause the neurons to become hyperpolarized. Norepinephrine has an inhibitory effect on MCH neurons as does acetylcholine. MCH neurons hyperpolarize in response to serotonin. Cannabinoids have an excitatory effect on MCH neurons.
Some research has shown that dopamine has an inhibitory effect on MCH neurons, but further research is needed to fully characterize this interaction.
Function.
Sleep.
MCH and the hormone orexin have an antagonistic relationship with one another with regards to the sleep cycle, with orexin being almost entirely active during wake periods and MCH being more active during sleep periods. MCH also promotes sleep, and within a sleep period increased levels of MCH seem to increase the amount of time spent in REM sleep and slow waves sleep. Increased levels of MCH can also increase the amount of time spent in both REM and NREM, which in turn increases total sleep duration. Increased levels of sugar promotes MCH and its effect on sleep and conserving energy.
Maternal behavior.
The presence of MCH in specific locations solely during lactation is thought to help to promote maternal behavior in individuals.
Eating behaviors and energy conservation.
An increased presence of MCH can cause increased eating levels and has been linked to an increase in body mass. Inversely, a decrease in the amount of MCH present can cause decreased levels in eating. Increased amounts of MCH in olfactory regions, among others, have also been linked to an increased intake of fatty foods with high caloric content. Food that is found to taste good also seems to promote MCH, which reinforces the eating of that food. Sugar, specifically glucose, seems to promote MCH's role in sleep and energy conservation. This promoting of energy conservation has also been linked to higher body mass even when diet is controlled.
Reproduction.
It has been postulated that MCH has a modulatory role with the release of Luteinizing Hormone (LH) either by directly acting on the pituitary gland or indirectly by affecting Gonadotropin-releasing hormone (GNRH) in the hypothalamus. Estrogen seems to be necessary in order for MCH to affect reproduction.
Skin pigmentation.
Though MCH was initially discovered for its role of determining the pigmentation in fish, trying to determine MCH's role in skin pigmentation in mammals has been much more difficult to determine. However, MCHR1 has been found in human melanocytes and some melanoma cells, so MCH is able to bind to these cells as well as keratinocytes though they do not express MCHR1. In melanocytes, MCH seems to have an antagonistic relationship with α-MSH, and decreased melanin production. Though, more information is needed to fully understand MCH's relationship with skin pigmentation in mammals.
Clinical significance.
Narcolepsy.
While MCH does promote sleep, there has been no research that links MCH to narcolepsy. Research has instead found that in individuals with narcolepsy there is a decrease in orexin neurons, which would promote wakefulness, while the number of MCH neurons do not vary from the average non-narcoleptic individual.
Depression and anxiety.
MCH has been linked to depression and anxiety. MCHR1 antagonists have been show to act as antidepressants.
Anorexia.
Interactions between MCH and chemokines/cytokines that lead to an overall decrease in MCH release and neuron excitability has been linked with infection-induced anorexia. Chemokines and cytokines often appear as the result of inflammation or infection, and they can then damage MCH neurons, which can lead to anorexia in an individual.
Skin cancers.
MCH has been identified in both melanoma and squamous cell carcinoma cell lines. However, pro-MCH, a precursor to MCH, has not been found in melanocytes, keratinocytes, or fibroblasts, which might indicate MCH might be brought into these cells by macrophages as part of the immune response. More research is needed to fully determine and understand any relationship between MCH and possible immune responses in skin.
History.
MCH was initially discovered in the teleost fish; it was found to help determine the fish's skin color. Later, a mammalian version of MCH was discovered in rats, where most of the functions and localizations are conserved across mammalian species.


2-Amino-4-hydroxy-6-pyrophosphoryl-methylpteridine

2-Amino-4-hydroxy-6-pyrophosphoryl-methylpteridine (7,8-Dihydropterin pyrophosphate, dihydropterin-CH2OH-diphosphate) is a pteridine; a precursor to dihydrofolic acid.


Isotachophoresis

Isotachophoresis (ITP) is a technique in analytical chemistry used for selective separation and concentration of ionic analytes. It is a form of electrophoresis; charged analytes are separated based on ionic mobility, a quantity which tells how fast an ion migrates through an electric field.
Overview.
In conventional ITP separations, a discontinuous buffer system is used. The sample is introduced between a zone of fast leading electrolyte (LE) and a zone of slow terminating (or: trailing) electrolyte (TE). Usually, the LE and the TE have a common counterion, but the co-ions (having charges with the same sign as the analytes of interest) are different: the LE is defined by co-ions with high ionic mobility, while the TE is defined by co-ions with low ionic mobility. The analytes of interest have intermediate ionic mobility. Application of an electric potential results in a low electrical field in the leading electrolyte and a high electrical field in the terminating electrolyte. Analyte ions situated in the TE zone will migrate faster than the surrounding TE co-ions, while analyte ions situated in the LE will migrate slower; the result is that analytes are focused at the LE/TE interface.
ITP is a displacement method: focusing ions of a certain kind displace other ions. If present in sufficient amounts, focusing analyte ions can displace all electrolyte co-ions, reaching a plateau concentration. Multiple analytes with sufficiently different ionic mobilities will form multiple plateau zones. Indeed, plateau mode ITP separations are readily recognized by stairlike profiles, each plateau of the stair representing an electrolyte or analyte zone having (from LE to TE) increasing electric fields and decreasing conductivities. In peak mode ITP, analytes amounts are insufficient to reach plateau concentrations, such analytes will focus in sharp Gaussian-like peaks. In peak mode ITP, analyte peaks will strongly overlap, unless so-called spacer compounds are added with intermediate ionic mobilities between those of the analytes; such spacer compounds are able to segregate adjacent analyte zones.
A completed ITP separation is characterized by a dynamic equilibrium in which all coionic zones migrate with equal velocities. From this phenomenon ITP has obtained its name: iso = equal, "tachos" = speed, "phoresis" = migration.
Isotachophoresis is exactly equal to the steady-state-stacking step in Discontinuous electrophoresis.
Transient ITP.
A popular form of ITP is transient ITP (tITP). It alleviates the limitation of conventional ITP that it has limited separation capacity because of analyte zone overlap. In transient ITP, analytes are first concentrated by ITP, and then can be baseline separated by zone electrophoresis. Transient ITP is usually accomplished by dissolving the sample in the TE and sandwiching the sample/TE plug between LE zones - or vice versa: a sample/LE plug can also be sandwiched between TE zones. In the first case, analytes are focused at the front TE/LE interface. Meanwhile, the back of the TE plug becomes dissolved in the LE because the faster LE ions overcome the TE ions. When all of the TE ions are dissolved, the focusing process ceases and the analytes are separated according to the principles of zone electrophoresis.
tITP is nowadays more widespread than conventional ITP because it is easily implemented in capillary electrophoresis (CE) separations as a preconcentration step, making CE more sensitive while profiting from its powerful separation capacities.


Minoru 3D Webcam

The Minoru 3D Webcam is a stereoscopic webcam produced by Promotion and Display Technology of Salford, Greater Manchester. Released in January 2009, it won the "Fans' Favorite" award at the Consumer Electronics Show, in Las Vegas. According to the company, it is the "world's first consumer 3D-webcam." It was designed by David Holder.
The webcam, which hooks to a computer via USB port, consists of two cameras, being "roughly the same distance apart as human eyes," that are held together in a device that resembles Wall-E, a Disney character, in appearance. It is capable of 3D, 2D, and Picture-in-picture graphics. Its image can be output from 320x240 pixels to 800x600 pixels, and is capable of 30 frames per second. The 3D imagery can be produced in anaglyph and side by side format.
The camera has two VGA 640x480 CMOS sensors, two high-quality wide-angle lens, and a built in USB microphone.
According to its website, it is most effective when the user is at a minimum of three feet away.
It derives its name from the Japanese term Minoru, which means "reality."


Thermal cycler

The thermal cycler (also known as a thermocycler, PCR machine or DNA amplifier) is a laboratory apparatus most commonly used to amplify segments of DNA via the polymerase chain reaction (PCR). Thermal cyclers may also be used in laboratories to facilitate other temperature-sensitive reactions, including restriction enzyme digestion or rapid diagnostics. The device has a "thermal block" with holes where tubes holding the reaction mixtures can be inserted. The cycler then raises and lowers the temperature of the block in discrete, pre-programmed steps.
History.
The earliest thermal cyclers were designed for use with the Klenow fragment of DNA polymerase I. Since this enzyme is destroyed during each heating step of the amplification process, new enzyme had to be added every cycle. This led to a cumbersome machine based on an automated pipettor, with open reaction tubes. Later, the PCR process was adapted to the use of thermostable DNA polymerase from "Thermus aquaticus", which greatly simplified the design of the thermal cycler. While in some old machines the block is submerged in an oil bath to control temperature, in modern PCR machines a Peltier element is commonly used. Quality thermal cyclers often contain silver blocks to achieve fast temperature changes and uniform temperature throughout the block. Other cyclers have multiple blocks with high heat capacity, each of which is kept at a constant temperature, and the reaction tubes are moved between them by means of an automated process. Miniaturized thermal cyclers have been created in which the reaction mixture moves via channel through hot and cold zones on a microfluidic chip. Thermal cyclers designed for quantitative PCR have optical systems which enable fluorescence to be monitored during reaction cycling.
Modern innovations.
Modern thermal cyclers are equipped with a heated lid that presses against the lids of the reaction tubes. This prevents condensation of water from the reaction mixtures on the insides of the lids. Traditionally, a layer of mineral oil was used for this purpose. Some thermal cyclers are equipped with a fully adjustable heated lid to allow for nonstandard or diverse types of PCR plasticware.
Some thermal cyclers are equipped with multiple blocks allowing several different PCRs to be carried out simultaneously. Some models also have a "gradient" function to allow for different temperatures in different parts of the block. This is particularly useful when testing suitable annealing temperatures for PCR primers.


Timberius


Sphaerozoum

Sphaerozoum is a radiolarian genus formerly reported in the subfamily Sphaerozoidae, now reported in the Collosphaeridae. The genus contains bioluminescent species. It is a genus of colonial radiolarians (as opposed to solitary).
Species.
The following species are recognized:


Steroid reductase

Steroid reductases are reductase enzymes that are involved in steroid biosynthesis and metabolism. They include:


Kareo

Kareo is an Irvine, California company that provides software as a service for independent medical practices. The company offers cloud computing products and services for electronic health record (EHR) management, medical practice management software, managed billing services and software to help practices engage with their patients. In 2019, the company reported over 55,000 providers using its technology.
History.
Kareo was established by former Scour Inc. founder Dan Rodrigues in 2004. The company is headquartered in Irvine, CA and has offices in Las Vegas, NV and Indianapolis, IN.
In July 2013, Kareo made its first acquisition of a full-service provider of medical billing and associated solutions company, ECCO Health, LLC.
Over the years, the company has received funding from a variety of venture capital firms including CNET founder Halsey Minor's Minor Ventures and OpenView Venture Partners.
On March 14, 2014, Kareo EHR achieved Meaningful Use 2014 Edition Stage 2 certification by the Drummond Group, which is an Office of the National Coordinator for Health Information Technology Authorized Certification Body (ONC-ACB).
Kareo acquired DoctorBase, a mobile-based patient engagement and practice marketing platform on March 10, 2015.
On April 2, 2015, the company announced that it joined Commonwell Health Alliance, a not-for-profit trade association of health IT companies working together to create universal access to health care data.
Awards.
In 2020, Kareo was ranked on the Investopedia "7 Best Medical Billing Companies" and rated as "Best for Independent Physician."


Intracrine

Intracrine refers to a hormone that acts inside a cell, regulating intracellular events. In simple terms it means that the cell stimulates itself by cellular production of a factor that acts within the cell. Steroid hormones act through intracellular (mostly nuclear) receptors and, thus, may be considered to be intracrines. In contrast, peptide or protein hormones, in general, act as endocrines, autocrines, or paracrines by binding to their receptors present on the cell surface. Several peptide/protein hormones or their isoforms also act inside the cell through different mechanisms. These peptide/protein hormones, which have intracellular functions, are also called intracrines. The term 'intracrine' is thought to have been coined to represent peptide/protein hormones that also have intracellular actions. To better understand intracrine, we can compare it to paracrine, autocrine and endocrine. The autocrine system deals with the autocrine receptors of a cell allowing for the hormones to bind, which have been secreted from that same cell. The paracrine system is one where nearby cells get hormones from a cell, and change the functioning of those nearby cells. The endocrine system refers to when the hormones from a cell affect another cell that is very distant from the one that released the hormone.
Paracrine physiology has been understood for decades now and the effects of paracrine hormones have been observed when for example, an obesity associate tumor will face the effects of local adipocytes, even if it is not in direct contact with the fat pads in concern. Endocrine physiology on the other hand is a growing field and has had a new area explored, called intracrinology. In intracrinology, the sex steroids produced locally, exert their action in the same cell where they are produced.
The biological effects produced by intracellular actions are referred as intracrine effects, whereas those produced by binding to cell surface receptors are called endocrine, autocrine, or paracrine effects, depending on the origin of the hormone. The intracrine effect of some of the peptide/protein hormones are similar to their endocrine, autocrine, or paracrine effects; however, these effects are different for some other hormones.
Intracrine can also refer to a hormone acting within the cell that synthesizes it.
Examples of intracrine peptide hormones: There are several protein/peptide hormones that are also intracrines. Notable examples that have been described in the references include:


Non-coding RNA

A non-coding RNA (ncRNA) is an RNA molecule that is not translated into a protein. The DNA sequence from which a functional non-coding RNA is transcribed is often called an RNA gene. Abundant and functionally important types of non-coding RNAs include transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs), as well as small RNAs such as microRNAs, siRNAs, piRNAs, snoRNAs, snRNAs, exRNAs, scaRNAs and the long ncRNAs such as Xist and HOTAIR.
The number of non-coding RNAs within the human genome is unknown; however, recent transcriptomic and bioinformatic studies suggest that there are thousands of them. Many of the newly identified ncRNAs have not been validated for their function. It is also likely that many ncRNAs are non functional (sometimes referred to as junk RNA), and are the product of spurious transcription.
Non-coding RNAs are thought to contribute to diseases including cancer and Alzheimer's.
History and discovery.
Nucleic acids were first discovered in 1868 by Friedrich Miescher and by 1939 RNA had been implicated in protein synthesis. Two decades later, Francis Crick predicted a functional RNA component which mediated translation; he reasoned that RNA is better suited to base-pair with an mRNA transcript than a pure polypeptide.
The first non-coding RNA to be characterised was an alanine tRNA found in baker's yeast, its structure was published in 1965. To produce a purified alanine tRNA sample, Robert W. Holley "et al." used 140kg of commercial baker's yeast to give just 1g of purified tRNAAla for analysis. The 80 nucleotide tRNA was sequenced by first being digested with Pancreatic ribonuclease (producing fragments ending in Cytosine or Uridine) and then with takadiastase ribonuclease Tl (producing fragments which finished with Guanosine). Chromatography and identification of the 5' and 3' ends then helped arrange the fragments to establish the RNA sequence. Of the three structures originally proposed for this tRNA, the 'cloverleaf' structure was independently proposed in several following publications. The cloverleaf secondary structure was finalised following X-ray crystallography analysis performed by two independent research groups in 1974.
Ribosomal RNA was next to be discovered, followed by URNA in the early 1980s. Since then, the discovery of new non-coding RNAs has continued with snoRNAs, Xist, CRISPR and many more. Recent notable additions include riboswitches and miRNA; the discovery of the RNAi mechanism associated with the latter earned Craig C. Mello and Andrew Fire the 2006 Nobel Prize in Physiology or Medicine.
Recent discoveries of ncRNAs have been achieved through both experimental and bioinformatic methods.
Biological roles.
Noncoding RNAs belong to several groups and are involved in many cellular processes. These range from ncRNAs of central importance that are conserved across all or most cellular life through to more transient ncRNAs specific to one or a few closely related species. The more conserved ncRNAs are thought to be molecular fossils or relics from the last universal common ancestor and the RNA world, and their current roles remain mostly in regulation of information flow from DNA to protein.
In translation.
Many of the conserved, essential and abundant ncRNAs are involved in translation. Ribonucleoprotein (RNP) particles called ribosomes are the 'factories' where translation takes place in the cell. The ribosome consists of more than 60% ribosomal RNA; these are made up of 3 ncRNAs in prokaryotes and 4 ncRNAs in eukaryotes. Ribosomal RNAs catalyse the translation of nucleotide sequences to protein. Another set of ncRNAs, Transfer RNAs, form an 'adaptor molecule' between mRNA and protein. The H/ACA box and C/D box snoRNAs are ncRNAs found in archaea and eukaryotes. RNase MRP is restricted to eukaryotes. Both groups of ncRNA are involved in the maturation of rRNA. The snoRNAs guide covalent modifications of rRNA, tRNA and snRNAs; RNase MRP cleaves the internal transcribed spacer 1 between 18S and 5.8S rRNAs. The ubiquitous ncRNA, RNase P, is an evolutionary relative of RNase MRP. RNase P matures tRNA sequences by generating mature 5'-ends of tRNAs through cleaving the 5'-leader elements of precursor-tRNAs. Another ubiquitous RNP called SRP recognizes and transports specific nascent proteins to the endoplasmic reticulum in eukaryotes and the plasma membrane in prokaryotes. In bacteria Transfer-messenger RNA (tmRNA) is an RNP involved in rescuing stalled ribosomes, tagging incomplete polypeptides and promoting the degradation of aberrant mRNA.
In RNA splicing.
In eukaryotes the spliceosome performs the splicing reactions essential for removing intron sequences, this process is required for the formation of mature mRNA. The spliceosome is another RNP often also known as the snRNP or tri-snRNP. There are two different forms of the spliceosome, the major and minor forms. The ncRNA components of the major spliceosome are U1, U2, U4, U5, and U6. The ncRNA components of the minor spliceosome are U11, U12, U5, U4atac and U6atac.
Another group of introns can catalyse their own removal from host transcripts; these are called self-splicing RNAs. There are two main groups of self-splicing RNAs: group I catalytic intron and group II catalytic intron. These ncRNAs catalyze their own excision from mRNA, tRNA and rRNA precursors in a wide range of organisms.
In mammals it has been found that snoRNAs can also regulate the alternative splicing of mRNA, for example snoRNA HBII-52 regulates the splicing of serotonin receptor 2C.
In nematodes, the SmY ncRNA appears to be involved in mRNA trans-splicing.
In DNA replication.
Y RNAs are stem loops, necessary for DNA replication through interactions with chromatin and initiation proteins (including the origin recognition complex). They are also components of the Ro60 ribonucleoprotein particle which is a target of autoimmune antibodies in patients with systemic lupus erythematosus.
In gene regulation.
The expression of many thousands of genes are regulated by ncRNAs. This regulation can occur in trans or in cis. There is increasing evidence that a special type of ncRNAs called enhancer RNAs, transcribed from the enhancer region of a gene, act to promote gene expression.
Trans-acting.
In higher eukaryotes microRNAs regulate gene expression. A single miRNA can reduce the expression levels of hundreds of genes. The mechanism by which mature miRNA molecules act is through partial complementary to one or more messenger RNA (mRNA) molecules, generally in 3' UTRs. The main function of miRNAs is to down-regulate gene expression.
The ncRNA RNase P has also been shown to influence gene expression. In the human nucleus RNase P is required for the normal and efficient transcription of various ncRNAs transcribed by RNA polymerase III. These include tRNA, 5S rRNA, SRP RNA, and U6 snRNA genes. RNase P exerts its role in transcription through association with Pol III and chromatin of active tRNA and 5S rRNA genes.
It has been shown that 7SK RNA, a metazoan ncRNA, acts as a negative regulator of the RNA polymerase II elongation factor P-TEFb, and that this activity is influenced by stress response pathways.
The bacterial ncRNA, 6S RNA, specifically associates with RNA polymerase holoenzyme containing the sigma70 specificity factor. This interaction represses expression from a sigma70-dependent promoter during stationary phase.
Another bacterial ncRNA, OxyS RNA represses translation by binding to Shine-Dalgarno sequences thereby occluding ribosome binding. OxyS RNA is induced in response to oxidative stress in Escherichia coli.
The B2 RNA is a small noncoding RNA polymerase III transcript that represses mRNA transcription in response to heat shock in mouse
cells. B2 RNA inhibits transcription by binding to core Pol II. Through this interaction, B2 RNA assembles into preinitiation
complexes at the promoter and blocks RNA synthesis.
A recent study has shown that just the act of transcription of ncRNA sequence can have an influence on gene expression. RNA polymerase II transcription of ncRNAs is required for chromatin remodelling in the Schizosaccharomyces pombe. Chromatin is progressively converted to an open configuration, as several species of ncRNAs are transcribed.
Cis-acting.
A number of ncRNAs are embedded in the 5' UTRs (Untranslated Regions) of protein coding genes and influence their expression in various ways. For example, a riboswitch can directly bind a small target molecule; the binding of the target affects the gene's activity.
RNA leader sequences are found upstream of the first gene of amino acid biosynthetic operons. These RNA elements form one of two possible structures in regions encoding very short peptide sequences that are rich in the end product amino acid of the operon. A terminator structure forms when there is an excess of the regulatory amino acid and ribosome movement over the leader transcript is not impeded. When there is a deficiency of the charged tRNA of the regulatory amino acid the ribosome translating the leader peptide stalls and the antiterminator structure forms. This allows RNA polymerase to transcribe the operon. Known RNA leaders are Histidine operon leader, Leucine operon leader, Threonine operon leader and the Tryptophan operon leader.
Iron response elements (IRE) are bound by iron response proteins (IRP). The IRE is found in UTRs of various mRNAs whose products are involved in iron metabolism. When iron concentration is low, IRPs bind the ferritin mRNA IRE leading to translation repression.
Internal ribosome entry sites (IRES) are RNA structures that allow for translation initiation in the middle of a mRNA sequence as part of the process of protein synthesis.
In genome defense.
Piwi-interacting RNAs (piRNAs) expressed in
mammalian testes and somatic cells form RNA-protein complexes with Piwi proteins. These piRNA complexes (piRCs) have been linked to transcriptional gene silencing of retrotransposons and other genetic elements in germ line cells, particularly those in spermatogenesis.
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) are repeats found in the DNA of many bacteria and archaea. The repeats are separated by spacers of similar length. It has been demonstrated that these spacers can be derived from phage and subsequently help protect the cell from infection.
Chromosome structure.
Telomerase is an RNP enzyme that adds specific DNA sequence repeats ("TTAGGG" in vertebrates) to telomeric regions, which are found at the ends of eukaryotic chromosomes. The telomeres contain condensed DNA material, giving stability to the chromosomes. The enzyme is a reverse transcriptase that carries Telomerase RNA, which is used as a template when it elongates telomeres, which are shortened after each replication cycle.
Xist (X-inactive-specific transcript) is a long ncRNA gene on the X chromosome of the placental mammals that acts as major effector of the X chromosome inactivation process forming Barr bodies. An antisense RNA, Tsix, is a negative regulator of Xist. X chromosomes lacking Tsix expression (and thus having high levels of Xist transcription) are inactivated more frequently than normal chromosomes. In drosophilids, which also use an XY sex-determination system, the roX (RNA on the X) RNAs are involved in dosage compensation. Both Xist and roX operate by epigenetic regulation of transcription through the recruitment of histone-modifying enzymes.
Bifunctional RNA.
"Bifunctional RNAs", or "dual-function RNAs", are RNAs that have two distinct functions. The majority of the known bifunctional RNAs are mRNAs that encode both a protein and ncRNAs. However, a growing number of ncRNAs fall into two different ncRNA categories; e.g., H/ACA box snoRNA and miRNA.
Two well known examples of bifunctional RNAs are SgrS RNA and RNAIII. However, a handful of other bifunctional RNAs are known to exist (e.g., steroid receptor activator/SRA, VegT RNA,
Oskar RNA, ENOD40, p53 RNA and SR1 RNA. Bifunctional RNAs have recently been the subject of a special issue of Biochimie.
As a hormone.
There is an important link between certain non-coding RNAs and the control of hormone-regulated pathways. In "Drosophila", hormones such as ecdysone and juvenile hormone can promote the expression of certain miRNAs. Furthermore, this regulation occurs at distinct temporal points within "C. elegans" development. In mammals, miR-206 is a crucial regulator of estrogen-receptor-alpha.
Non-coding RNAs are crucial in the development of several endocrine organs, as well as in endocrine diseases such as diabetes mellitus. Specifically in the MCF-7 cell line, addition of 17β-estradiol increased global transcription of the noncoding RNAs called lncRNAs near estrogen-activated coding genes.
In pathogenic avoidance.
"C. elegans" was shown to learn and inherit pathogenic avoidance after exposure to a single non-coding RNA of a bacterial pathogen.
Roles in disease.
As with proteins, mutations or imbalances in the ncRNA repertoire within the body can cause a variety of diseases.
Cancer.
Many ncRNAs show abnormal expression patterns in cancerous tissues. These include miRNAs, long mRNA-like ncRNAs, GAS5, SNORD50, telomerase RNA and Y RNAs. The miRNAs are involved in the large scale regulation of many protein coding genes, the Y RNAs are important for the initiation of DNA replication, telomerase RNA that serves as a primer for telomerase, an RNP that extends telomeric regions at chromosome ends (see telomeres and disease for more information). The direct function of the long mRNA-like ncRNAs is less clear.
Germ-line mutations in miR-16-1 and miR-15 primary precursors have been shown to be much more frequent in patients with chronic lymphocytic leukemia compared to control populations.
It has been suggested that a rare SNP (rs11614913) that overlaps has-mir-196a2 has been found to be associated with non-small cell lung carcinoma. Likewise, a screen of 17 miRNAs that have been predicted to regulate a number of breast cancer associated genes found variations in the microRNAs miR-17 and miR-30c-1of patients; these patients were noncarriers of BRCA1 or BRCA2 mutations, lending the possibility that familial breast cancer may be caused by variation in these miRNAs.
The p53 tumor suppressor is arguably the most important agent in preventing tumor formation and progression. The p53 protein functions as a transcription factor with a crucial role in orchestrating the cellular stress response. In addition to its crucial role in cancer, p53 has been implicated in other diseases including diabetes, cell death after ischemia, and various neurodegenerative diseases such as Huntington, Parkinson, and Alzheimer. Studies have suggested that p53 expression is subject to regulation by non-coding RNA.
Another example of non-coding RNA dysregulated in cancer cells is the long non-coding RNA Linc00707. Linc00707 is upregulated and sponges miRNAs in human bone marrow-derived mesenchymal stem cells, in hepatocellular carcinoma, gastric cancer or breast cancer, and thus promotes osteogenesis, contributes to hepatocellular carcinoma progression, promotes proliferation and metastasis, or indirectly regulates expression of proteins involved in cancer aggressiveness, respectively.
Prader–Willi syndrome.
The deletion of the 48 copies of the C/D box snoRNA SNORD116 has been shown to be the primary cause of Prader–Willi syndrome. Prader–Willi is a developmental disorder associated with over-eating and learning difficulties. SNORD116 has potential target sites within a number of protein-coding genes, and could have a role in regulating alternative splicing.
Autism.
The chromosomal locus containing the small nucleolar RNA SNORD115 gene cluster has been duplicated in approximately 5% of individuals with autistic traits. A mouse model engineered to have a duplication of the SNORD115 cluster displays autistic-like behaviour. A recent small study of post-mortem brain tissue demonstrated altered expression of long non-coding RNAs in the prefrontal cortex and cerebellum of autistic brains as compared to controls.
Cartilage–hair hypoplasia.
Mutations within RNase MRP have been shown to cause cartilage–hair hypoplasia, a disease associated with an array of symptoms such as short stature, sparse hair, skeletal abnormalities and a suppressed immune system that is frequent among Amish and Finnish. The best characterised variant is an A-to-G transition at nucleotide 70 that is in a loop region two bases 5' of a conserved pseudoknot. However, many other mutations within RNase MRP also cause CHH.
Alzheimer's disease.
The antisense RNA, BACE1-AS is transcribed from the opposite strand to BACE1 and is upregulated in patients with Alzheimer's disease. BACE1-AS regulates the expression of BACE1 by increasing BACE1 mRNA stability and generating additional BACE1 through a post-transcriptional feed-forward mechanism. By the same mechanism it also raises concentrations of beta amyloid, the main constituent of senile plaques. BACE1-AS concentrations are elevated in subjects with Alzheimer's disease and in amyloid precursor protein transgenic mice.
miR-96 and hearing loss.
Variation within the seed region of mature miR-96 has been associated with autosomal dominant, progressive hearing loss in humans and mice. The homozygous mutant mice were profoundly deaf, showing no cochlear responses. Heterozygous mice and humans progressively lose the ability to hear.
Distinction between functional RNA (fRNA) and ncRNA.
Scientists have started to distinguish "functional RNA" ("fRNA") from ncRNA, to describe regions functional at the RNA level that may or may not be stand-alone RNA transcripts. This implies that fRNA (such as riboswitches, SECIS elements, and other cis-regulatory regions) is not ncRNA. Yet fRNA could also include mRNA, as this is RNA coding for protein, and hence is functional. Additionally artificially evolved RNAs also fall under the fRNA umbrella term. Some publications state that "ncRNA" and "fRNA" are nearly synonymous, however others have pointed out that a large proportion of annotated ncRNAs likely have no function. It also has been suggested to simply use the term "RNA", since the distinction from a protein coding RNA (messenger RNA) is already given by the qualifier "mRNA". This eliminates the ambiguity when addressing a gene "encoding a non-coding" RNA. Besides, there may be a number of ncRNAs that are misannoted in published literature and datasets.


James Sherley

James Sherley is a biological engineer and the founder of Asymmetrex, an adult stem cell research center. He has also conducted research at the Boston Biomedical Research Institute and Massachusetts Institute of Technology (MIT). Sherley filed a suit against the government in "Sherley v. Sebelius", resulting in a protracted legal battle attempting to ban the government from funding any research relating to embryonic stem cells.
Early life and education.
Sherley's education includes a B.S. from Harvard University and an M.D and a Ph.D from Johns Hopkins University School of Medicine, Baltimore, Maryland. He was a Postdoctoral Fellow with Arnold J. Levine, at Princeton University, Princeton, New Jersey 1988 to 1991.
From 1991 to 1998 he was Associate Member, Division of Medical Science, Molecular Oncology Group, Fox Chase Cancer Center, Philadelphia, Pennsylvania. He was a professor in Biological Engineering Division at Massachusetts Institute of Technology from 1998 to 2007.
Career.
His awards include a 1993 Pew Scholar award and recipient of a 2006 NIH Director's Pioneer Award.
In December 2006, Sherley announced he would protest MIT's decision to not grant him tenure by going on a hunger strike. He ended the strike 12 days later. He subsequently asserted that he would continue to show up for work on July 1, 2007, despite no longer having a job at MIT on that date. Twenty senior faculty members who participated in evaluation of his tenure case issued a public statement saying that Sherley's evaluation was conducted with integrity and that he was treated fairly.
Sherley, an African-American, was one of 23 black professors at MIT. As of October 2006, of the 988 faculty members, 165 or about 17% are ethnic minorities. He has stated that he believes that MIT has not given him the freedom to challenge scientific orthodoxy the way the institution would have for a white colleague. Some other colleagues and professors have defended Sherley, including MIT linguistics professor Noam Chomsky and Harvard medical school's George M. Church. Frank L. Douglas, then the director of the Center for Biomedical Innovation, resigned from the university as a result of the handling of the case.
Stem cell research.
He is open about his opposition to human embryonic stem cell research, and instead, Sherley's research focuses on the molecular and biochemical mechanisms of adult stem cells. In 2006 he stated that cloning human embryos was wrong and that the research from embryonic stem cell research would not lead to a cure all. "Despite similar misinformation to the contrary, adult stem cell research is a viable and vibrant path to new medical therapies. Even calling them an alternative to embryonic stem cells misinforms the public."
In 2010, Sherley was the named plaintiff in "Sherley v. Sebelius" against Kathleen Sebelius, director of HHS, and Francis Collins, director of the NIH, a case challenging federal (NIH) funding of embryonic stem cell research that resulted in a suspension of much of the NIH funded research on embryonic stem cells. He fought to ban the NIH from funding any research related to embryonic stem cells, and claimed that the decision of the federal government to pay for research on embryonic stem cells was "essentially robbing him of grant money by increasing the competition for funding". After three years of legal effort, the supreme court refused to overturn a lower judge's ruling against Sherley, allowing research to proceed.
Sherley has likened abortion to racism, claiming that it is "discriminating against human embryos, just like there is discrimination against people of different culture and races".
While working at the Boston Biomedical Research Institute in 2009, Sherley founded a stem cell research center known as Asymmetrex.


Diethyl lutidinate

Diethyl lutidinate is a chemical compound. It has been studied for its potential use in hair care.


Northern blot

The northern blot, or RNA blot, is a technique used in molecular biology research to study gene expression by detection of RNA (or isolated mRNA) in a sample.
With northern blotting it is possible to observe cellular control over structure and function by determining the particular gene expression rates during differentiation and morphogenesis, as well as in abnormal or diseased conditions. Northern blotting involves the use of electrophoresis to separate RNA samples by size, and detection with a hybridization probe complementary to part of or the entire target sequence. The term 'northern blot' actually refers specifically to the capillary transfer of RNA from the electrophoresis gel to the blotting membrane. However, the entire process is commonly referred to as northern blotting. The northern blot technique was developed in 1977 by James Alwine, David Kemp, and George Stark at Stanford University, with contributions from Gerhard Heinrich. Northern blotting takes its name from its similarity to the first blotting technique, the Southern blot, named for biologist Edwin Southern. The major difference is that RNA, rather than DNA, is analyzed in the northern blot.
Procedure.
A general blotting procedure starts with extraction of total RNA from a homogenized tissue sample or from cells. Eukaryotic mRNA can then be isolated through the use of oligo (dT) cellulose chromatography to isolate only those RNAs with a poly(A) tail. RNA samples are then separated by gel electrophoresis. Since the gels are fragile and the probes are unable to enter the matrix, the RNA samples, now separated by size, are transferred to a nylon membrane through a capillary or vacuum blotting system. A nylon membrane with a positive charge is the most effective for use in northern blotting since the negatively charged nucleic acids have a high affinity for them. The transfer buffer used for the blotting usually contains formamide because it lowers the annealing temperature of the probe-RNA interaction, thus eliminating the need for high temperatures, which could cause RNA degradation. Once the RNA has been transferred to the membrane, it is immobilized through covalent linkage to the membrane by UV light or heat. After a probe has been labeled, it is hybridized to the RNA on the membrane. Experimental conditions that can affect the efficiency and specificity of hybridization include ionic strength, viscosity, duplex length, mismatched base pairs, and base composition. The membrane is washed to ensure that the probe has bound specifically and to prevent background signals from arising. The hybrid signals are then detected by X-ray film and can be quantified by densitometry. To create controls for comparison in a northern blot, samples not displaying the gene product of interest can be used after determination by microarrays or RT-PCR.
Gels.
The RNA samples are most commonly separated on agarose gels containing formaldehyde as a denaturing agent for the RNA to limit secondary structure. The gels can be stained with ethidium bromide (EtBr) and viewed under UV light to observe the quality and quantity of RNA before blotting. Polyacrylamide gel electrophoresis with urea can also be used in RNA separation but it is most commonly used for fragmented RNA or microRNAs. An RNA ladder is often run alongside the samples on an electrophoresis gel to observe the size of fragments obtained but in total RNA samples the ribosomal subunits can act as size markers. Since the large ribosomal subunit is 28S (approximately 5kb) and the small ribosomal subunit is 18S (approximately 2kb) two prominent bands appear on the gel, the larger at close to twice the intensity of the smaller.
Probes.
Probes for northern blotting are composed of nucleic acids with a complementary sequence to all or part of the RNA of interest, they can be DNA, RNA, or oligonucleotides with a minimum of 25 complementary bases to the target sequence. RNA probes (riboprobes) that are transcribed in vitro are able to withstand more rigorous washing steps preventing some of the background noise. Commonly cDNA is created with labelled primers for the RNA sequence of interest to act as the probe in the northern blot. The probes must be labelled either with radioactive isotopes (32P) or with chemiluminescence in which alkaline phosphatase or horseradish peroxidase (HRP) break down chemiluminescent substrates producing a detectable emission of light. The chemiluminescent labelling can occur in two ways: either the probe is attached to the enzyme, or the probe is labelled with a ligand (e.g. biotin) for which the ligand (e.g., avidin or streptavidin) is attached to the enzyme (e.g. HRP). X-ray film can detect both the radioactive and chemiluminescent signals and many researchers prefer the chemiluminescent signals because they are faster, more sensitive, and reduce the health hazards that go along with radioactive labels. The same membrane can be probed up to five times without a significant loss of the target RNA.
Applications.
Northern blotting allows one to observe a particular gene's expression pattern between tissues, organs, developmental stages, environmental stress levels, pathogen infection, and over the course of treatment. The technique has been used to show overexpression of oncogenes and downregulation of tumor-suppressor genes in cancerous cells when compared to 'normal' tissue, as well as the gene expression in the rejection of transplanted organs. If an upregulated gene is observed by an abundance of mRNA on the northern blot the sample can then be sequenced to determine if the gene is known to researchers or if it is a novel finding. The expression patterns obtained under given conditions can provide insight into the function of that gene. Since the RNA is first separated by size, if only one probe type is used variance in the level of each band on the membrane can provide insight into the size of the product, suggesting alternative splice products of the same gene or repetitive sequence motifs. The variance in size of a gene product can also indicate deletions or errors in transcript processing. By altering the probe target used along the known sequence it is possible to determine which region of the RNA is missing.
Advantages and disadvantages.
Analysis of gene expression can be done by several different methods including RT-PCR, RNase protection assays, microarrays, RNA-Seq, serial analysis of gene expression (SAGE), as well as northern blotting. Microarrays are quite commonly used and are usually consistent with data obtained from northern blots; however, at times northern blotting is able to detect small changes in gene expression that microarrays cannot. The advantage that microarrays have over northern blots is that thousands of genes can be visualized at a time, while northern blotting is usually looking at one or a small number of genes.
A problem in northern blotting is often sample degradation by RNases (both endogenous to the sample and through environmental contamination), which can be avoided by proper sterilization of glassware and the use of RNase inhibitors such as DEPC (diethylpyrocarbonate). The chemicals used in most northern blots can be a risk to the researcher, since formaldehyde, radioactive material, ethidium bromide, DEPC, and UV light are all harmful under certain exposures. Compared to RT-PCR, northern blotting has a low sensitivity, but it also has a high specificity, which is important to reduce false positive results.
The advantages of using northern blotting include the detection of RNA size, the observation of alternate splice products, the use of probes with partial homology, the quality and quantity of RNA can be measured on the gel prior to blotting, and the membranes can be stored and reprobed for years after blotting.
For northern blotting for the detection of acetylcholinesterase mRNA the nonradioactive technique was compared to a radioactive technique and found as sensitive as the radioactive one, but requires no protection against radiation and is less time consuming.
Reverse northern blot.
Researchers occasionally use a variant of the procedure known as the reverse northern blot. In this procedure, the substrate nucleic acid (that is affixed to the membrane) is a collection of isolated DNA fragments, and the probe is RNA extracted from a tissue and radioactively labelled.
The use of DNA microarrays that have come into widespread use in the late 1990s and early 2000s is more akin to the reverse procedure, in that they involve the use of isolated DNA fragments affixed to a substrate, and hybridization with a probe made from cellular RNA. Thus the reverse procedure, though originally uncommon, enabled northern analysis to evolve into gene expression profiling, in which many (possibly all) of the genes in an organism may have their expression monitored.


Thomas Hele (academic)

Thomas Shirley Hele, OBE, MD, FRCP (b Carlisle 24 October 1881 - d Cambridge 23 January 1953) was an academic in the 20th century.
Hele was educated at Carlisle Grammar School ; Sedbergh School; Emmanuel College, Cambridge (Fellow, 1911); and Barts. He was University Lecturer in Biochemistry from 1921; Tutor at Emmanuel from 1922 to 1935; its Master from 1935 to 1951; and Vice-Chancellor of the University of Cambridge from 1943 to 1945.


HBV RNA encapsidation signal epsilon

The HBV RNA encapsidation signal epsilon (HBV_epsilon) is an element essential for HBV virus replication.
It is an RNA structure situated near the 5' end of the HBV pregenomic RNA.
The structure consists of a lower stem, a bulge region, an upper stem and a tri-loop.
The structure was determined and refined through enzymatic probing and NMR spectroscopy.
The closure of the tri-loop was not predicted by RNA structure prediction programs but observed in the NMR structure.
The regions shown to be critical for encapsidation of the RNA in the viral lifecycle are the bulge, upper stem and tri-loop which interact with the terminal protein domain of the HBV viral polymerase.


David Landsborough Thomson

David Landsborough Thomson F.R.S.C., (1901 - 1964) was a Canadian biochemist, best known for the co-discovery of Adrenocorticotropic hormone (adreno-cortical thyroid hormone or ACTH) and as the vice-principal of McGill University. ACTH was co-discovered by Evelyn M. Anderson, James Bertram Collip and Thomson. In a paper published in 1933, they explained its function in the body.
Born in Scotland, Thomson earned BSc and MA degrees from the University of Aberdeen, then a PhD in biochemistry from Cambridge University under the eye of Nobel laureate Frederick Gowland Hopkins. After further studies in Europe, he moved to Montreal, joining the McGill faculty in 1928.
At McGill, he was Gilman Cheney Professor of Biochemistry from 1937 to 1960, dean of the faculty of graduate studies and research from 1942 to 1963, and the vice-principal from 1955 to 1963. He served on the National Research Council of Canada, the Defence Research Board and the Scientific Research Bureau of Quebec. He received an honorary LL.D. in 1961 from the University of Saskatchewan.
Like all British subjects then living in Canada, he became a Canadian with the passage of the Canadian Citizenship Act 1946.
Publications.
Thomson published several highly cited papers in major journals, including The Lancet, BMJ, NATURE, CMAJ, and J. Physiol. Many of these were in collaboration with Collip, Hans Selye, or both. Topic areas included endocrinology (particularly antihormones), vitamins (he coined the term "Vitamin B complex" in a 1938 paper),


PTD-DBM

Protein Transduction Domain-fused Dishevelled Binding Motif (PTD-DBM) is a man-made peptide which interacts with the mechanism of the hair loss linked endogenous protein, "CXXC5", which is a negative feedback regulator of the Wnt/β-catenin pathway"." Application of the peptide to bald laboratory mice resulted in new hair follicle growth.
PTD-DBM is a peptide activating the Wnt/β-catenin signaling pathway functioning via interference of the binding of CXXC5 to Dishevelled (Dvl), an upstream component of the Wnt/β-catenin pathway. By topical application, the PTD-DBM promotes the formation of new hair follicles and prevents hair loss. Combinatory treatment of PTD-DBM with valproic acid (VPA), the activator of Wnt/β-catenin pathway, further induce hair re-growth as well as wound-induced hair neogenesis (WIHN). The increased expression of CXXC5 in the bald scalps and excellent effects of PTD-DBM on hair growth in mice raised hopes for the application of this peptide on hair growth in the clinic.
Discovery of PTD-DBM.
Professor Kang-Yell Choi and his research team at Yonsei University in South Korea discovered a protein responsible for hair loss in the condition known as androgenetic alopecia. The responsible protein is called CXXC-type zinc finger protein 5 (CXXC5), which acts as a negative regulator for the Wnt/β-catenin pathway, involved in hair regeneration and wound healing. CXXC5 negatively regulates hair growth, and the researchers developed a new substance that promotes hair regeneration by controlling the function of CXXC5. When CXXC5 binds with the Dvl protein, which functions at the upstream of Wnt/β-catenin pathway, it suppresses hair regrowth and hair follicle neogenesis. The observation of CXXC5 overexpression in the bald scalp by Professor Choi’s team led to the development of PTD-DBM, which interferes with the CXXC5-Dvl protein-protein interaction (PPI). By topical application, PTD-DBM enhances hair regrowth as well as neogenesis. The hair growth promoting effect of PTD-DBM is further enhanced when used in combination with a Wnt/β-catenin signaling activator such as VPA, which is generally used as a drug for bipolar disorder and activates the Wnt/β-catenin pathway by inhibition of GSK3β. Currently, topical application of PTD-DBM or its combination with VPA has been used for treatment of hair loss.