Cell Science Exam Topic List PDF

Vishal Kumar CELL SCIENCE – EXAM TOPIC LIST 0 W i e b Vi hal K ma 2019/2020 Vishal Kumar. ORIGIN OF LIFE​ ​ ​ lecture of DGCI​ st​ Urey- Miller experiment In , Stanley Miller and Harold Urey did an...

Vishal Kumar CELL SCIENCE – EXAM TOPIC LIST 0 W i e b Vi hal K ma 2019/2020 Vishal Kumar. ORIGIN OF LIFE​ ​ ​ lecture of DGCI​ st​ Urey- Miller experiment In , Stanley Miller and Harold Urey did an experiment to test Oparin and Haldane’s ideas. Miller and Urey built a closed system containing a heated pool of water and a mixture of gases that were thought to be abundant in the atmosphere of early earth H​ O, ​ NH​ ,​ CH​ ,​ N​ ​. To simulate the lightning that might have provided energy for chemical reactions in Earth’s early atmosphere, Miller and Urey sent sparks of electricity through their experimental system. After letting the experiment run for a week, Miller and Urey found that various types of amino acids, sugars, lipids and other organic molecules had formed. Large, complex molecules like DNA and protein were missing, but the Miller-Urey experiment showed that at least ​some​ of the building blocks for these molecules could form spontaneously from simple compounds. Steps of chemical and biological evolution CHEMICAL EVOLUTION: Solidification of the Earth crust, development of the atmosphere formation of small organic molecules formation of large organic molecules formation of protocells BIOLOGICAL EVOLUTION: Prokaryote cells eukaryote single cell organisms multicellular organisms Three domains of cellular life Bacteria, Archea and Eukaryotes archaebacterial chimeras LUCA last universal common ancestor. S rRNA, a molecule conserved in all cellular life forms. Vishal Kumar. E​ ukaryotic cell organization and medicinal model cells.​ ​ ​ECB​: OR ​ - ​- very basic morphological background knowledge, , OR ​ -​ ​ , - ​- basic chemical protein background knowledge, ​chemical formulas are not needed​. ​ ​ lecture of DGCI​ st​ Main differences between pro- and eukaryotic cells The main difference is that eukaryotic cells have a nucleus, while the DNA of prokaryotic cells floats freely inside the cell. Eukaryotes have a plasma membrane, while prokaryotic have a cell wall with an outer membrane and an inner plasma membrane. Also, the bigger eukaryote contains several organelles which makes it more complex, which again gives it more ability to develop and adapt, and not least grow. Components of eukaryotic cell Components of eukaryotic cell: Plasma membrane Surrounds the cell. Selective permeability. cell membrane Cytosol part of cytoplasm and has a gel-like consistence because of the high concentration of molecules. Contains ribosomes. ( intracellar liquid ) Golgi complex Receives and “edits” molecules made in ER. Directs them to the exterior of the cell to various locations. Rough endoplasmic reticulum Contains ribosomes. Thus, proteins are synthesized here. Smooth endoplasmic reticulum Membrane closed maze-like structure, where most of the cell membrane parts as well as materials destined for export from the cell are made. Also functions as a detoxification site. Ribosome Where the translation from tRNA to proteins occur. Mitochondrion The cell´s powerhouse. It is believed that they once were prokaryotic cells which were taken up by endocytosis into eukaryotic cells. Their task is to make ATP mechanic energy from chemical energy food. Without them, the body would be anaerobic. Centrosome A centrosome contains a pair of centrioles. Functions as a main organizer of microtubules. Therefore, it plays an important role during mitosis, when the centrioles migrate to either horizontal end of the cell nucleus and form the mitotic spindle which pulls the sister chromatids away from each other. Vishal Kumar Lysosome Intracellular digestion. Microvilli Like small strands of hair, that increase surface area, which gives the cell various organelles higher absorption rate. Consists of simple squamous/cuboidal epithelial Increase absorption further. Cytoskeleton Responsible for structure of cell. Important role in Mitosis, by splitting sister chromatids microtubule. Consists of microtubules, actin filaments and intermediate filaments. Microtubule Thickest filaments that are part of cytoskeleton. Thus, they are essential in cell structure, and they also play a role in movement of vesicles and organelles inside the cell. Preform the splitting of sister chromatids in mitosis, through forming mitotic spindle. Peroxisome Where hydrogen peroxide is generated and degraded. In other words, breakes down toxic chemicals Nucleus nuclear envelope Contains the nucleolus. Protection of the DNA. Vishal Kumar Main biological macromolecules: carbohydrates, lipids, proteins, nucleic acids, and their monomers Basic compounds of cell Sugars Polysaccharides Fatty acids Fats, Lipids Amino acids Proteins : number and kinds Nucleotides Nucleic acids optical isomers same speilbilde) of atoms bonds , but ( and different spatial arrangement Sugars The simplest sugars, monosaccharides glucose C​ ​H​ ​O​ ​ carry many special properties which make them essential to cells. By switching the orientation of a OH group glucose can be converted into a whole new molecule. Furthermore, monosaccharides can exist in two & H shapes called the D-form and L-form, which are mirror images of each other, which makes o = , # o H H - them optical isomers. This plays a big role in the large variety of sugars which can exist. H H H H By linking two monosaccharides by covalent bonds called glyosidic bonds , a disaccharide is ( O H Hz formed. Even more linked together make up the polysaccharides. This is called D H carbohydrates. This linking happens through a condensation reaction between two OH o = , # H OH - groups, where a water molecule is expelled of. The reverse reaction is hydrolysis, where a H H water molecule is consumed. H H ( O H Hz The main function of sugars is to be provide energy by being broken down in the cell. But also, they can provide a structural/mechanical support cellulose. L the Ott the sealer circle is left - on D - the OH on the chiral center is right. Fatty acids A fatty acid consists of two major parts head tail. TAIL: One long hydrocarbon chain which is hydrophobic, and not very chemical reactive. HEAD: The other is a carboxyl COOH group which behaves as an acid. It is hydrophilic. Because the molecule is both hydrophilic and hydrophobic it is considered amphipathic. Furthermore, a fatty acid can be saturated or unsaturated, both respectively giving different properties to the molecule. ​Sat rated​ no not double bonds, allows molecule to be packed tightly together. Solid at room temperature. healthy. Unsat rated​ Double bonds create bends in the tail and prevents the molecule from packing tightly. Liquid at room temperature. Lipids are insoluble in water, but soluble in fat and organic solvents. Lipids function as an energy reserve for the human body. In the body, they are best known for constructing the plasma membrane which covers all cells; phospholipids. Since there would be no life without this membrane, lipids are essential for life. Vishal Kumar Amino Acids Amino acids are the building blocks of proteins. There are amino acids, and they all contain an amino group, a carboxyl group, and a different R-group which is individual for every amino acid. They are all connected to the same “alfa” carbon atom. All amino acids have different properties, and this is an important factor to why proteins are so complex. When two amino acids come together, they form a peptide bond. A chain of amino acids is known as a polypeptide. Peptide bonds are formed by condensation reactions. Nucleotides Nucleotides consists of nucleosides. Nucleosides are molecules made of a nitrogen-containing ring compound linked to a five-carbon sugar, which can be ribose or deoxyribose. A nucleoside containing one or more phosphate groups attached to its sugar is called a nucleotide. Nucleotides can act as short-term carriers of chemical energy. Adenosine triphosphate ATP is a nucleotide. Its three phosphates are linked in series by two phosphoanhydride bonds, which release large amount of useful energy when cleaved off. RNA contains A, G, C and U, while DNA contain A, G, C and T. Pyrimidines -ring : Cytosine, thymine and uracil. Purine -ring : Guanine and adenine. U and T have the same properties. RNA is a short-term on stranded information carrier, while the double stranded DNA stores the information. Macromolecules Macromolecules have in common that they are all built by adding subunits sugars, amino acids and nucleotides to the end of a strand by condensation expelling one water molecule. Also, this happens in ​sequence​, which means in a fixed order. Subunits form covalent bonds with each other, which creates macromolecules. Macromolecules form weak noncovalent bonds which play a huge role in which properties the molecules have. It also forms new structures and decides which macromolecules that can bond to form macromolecular complexes, such as ribosomes. Vishal Kumar Model cells (etc. HeLa, stem cells) Cells enabling studies on the structure or function of an organ or a tissue. Data obtained with model cells may serve as reference in: fundamental science, clinical dignostics or drug development. The benefits of using model cells: - they enable studies under standard laboratory conditions - the experimental conditions can be easily modified - high number of model cells can be investigated enabling statistical analysis - the use of model cells may save the lives of many experimental animals Eserichia Coli E.Coli is an example of model cell. Easy to grow in the lab, can be grown into millions of copies and easily manipulated genetically. Vishal Kumar. Structure and functions of plasma membrane​ ​ ​ECB​: - OR ​ – 0, - , 0 - 0 ​. ​ st lecture of DGCI​ Types and the role of lipids, proteins and carbohydrates in the membranes The principal components of a plasma membrane are lipids phospholipids and cholesterol , proteins, and carbohydrates attached to some of the lipids and some of the proteins. Lipids:​ The main fabric of the membrane is composed of amphiphilic phospholipid molecules. The hydrophilic or water-loving areas of these molecules are in contact with the aqueous fluid both inside and outside the cell. Hydrophobic, or water-hating molecules, tend to be non- polar. A phospholipid molecule consists of a three-carbon glycerol backbone with two fatty acid molecules attached to carbons and , and a phosphate-containing group attached to the third carbon. This arrangement gives the overall molecule an area described as its head the phosphate-containing group , which has a polar character or negative charge, and an area called the tail the fatty acids , which has no charge. When placed in water, hydrophobic molecules tend to form a ball or cluster. The hydrophilic regions of the phospholipids tend to form hydrogen bonds with water and other polar molecules on both the exterior and interior of the cell. Thus, the membrane surfaces that face the interior and exterior of the cell are hydrophilic. In contrast, the middle of the cell membrane is hydrophobic and will not interact with water. Therefore, phospholipids form an excellent lipid bilayer cell membrane that separates fluid within the cell from the fluid outside of the cell. Proteins: ​Proteins make up the second major component of plasma membranes. Integral proteins some specialized types are called integrins are, as their name suggests, integrated completely into the membrane structure, and their hydrophobic membrane-spanning regions interact with the hydrophobic region of the phospholipid bilayer. Some complex proteins are composed of up to segments of a single protein, which are ( innebygd ) extensively folded and embedded in the membrane. This type of protein has a hydrophilic region s , and one or several hydrophobic regions. This arrangement of regions of the protein tends to orient the protein alongside the phospholipids, with the hydrophobic region of the protein next to the tails of the phospholipids and the hydrophilic region s of the protein protruding from the membrane and in contact with the cytosol or extracellular fluid. Carbohydrates​: Carbohydrates are the third major component of plasma membranes. They are always found on the exterior surface of cells and are bound either to proteins forming glycoproteins or to lipids forming glycolipids. These carbohydrate chains may consist of monosaccharide units and can be either straight or branched. Along with peripheral proteins, carbohydrates form specialized sites on the cell surface that allow cells to recognize each other. This recognition function is very important to cells, as it allows the  oar sites spesial other form that makeseach that Vishal Kumar membraneto Guice carbohydrates co re cell cell the phevipheral t +u p the immune system to differentiate between body cells called “self” and foreign cells or tissues called “non-self”. These carbohydrates on the exterior surface of the cell the carbohydrate components of both glycoproteins and glycolipids are collectively referred to as the glycocalyx meaning “sugar coating”. Fluid mosaic membrane model The fluid mosaic model is the most acceptable model of the plasma membrane. Its main function is to give shape to the cell. The fluid mosaic model of the plasma membrane describes the plasma membrane as a fluid combination of phospholipids, cholesterol, and proteins. Carbohydrates attached to lipids glycolipids and to proteins glycoproteins extend from the outward-facing surface of the membrane. Dynamism, fluidity, asymmetry, inhomogeneity of membranes Dynamics​:​ When a cell grows or changes shape, so does its membrane: it enlarges in area by adding new membrane without ever losing its continuity, and it can deform without tearing. Transbilayer diffusion or ‘’flip-flop’’: in this type of movement phospholipids either move from the outer leaflet to the inner leaflet or the opposite way. Since this movement happens without a catalyst, it is very slow and does not occur so often. Lateral diffusion: is a type of diffusion where phospholipids move side to side in one leaflet. Movement is not only side to side, but in all directions. Since the phospholipids do not switch between leaflets, this type of movement is very fast and occurs often. Uncatalyzed. Flippase: almost the same type of movement as transbilayer diffusion, only that this type is aided by a protein, so it is catalyzed. Phospholipid on the outer leaflet moves to the inner leaflet. This process also uses ATP; breaks down to ADP P. This provides energy for this reaction to happen. Fast movement. Floppase: same type as flippase, just that a phospholipid moves from inner leaflet to outer leaflet. Is also fast, since it is catalyzed. Vishal Kumar Scramblase: this type of movement is a combination of flippase and floppase; phospholipids go from one leaflet to another in both directions. But this type does not need ATP. Is also fast, since it is catalyzed. Fluidity​: The fluidity of a cell membrane; how its lipid molecules can move in and out of the bilayer, is very important. Fluidity of a lipid membrane depends on its phospholipid composition, particularly, how close the packing of hydrocarbon tails is. At Celsius degree the plasma membrane is close to a fluid state A shorter chain length reduces the tendency of the hydrocarbon tails to interact with one another and therefore increases the fluidity of the bilayer. The closer and more regular the packing of tails, the more viscous and less fluid the bilayer will be. Most phospholipids contain one hydrocarbon tail that has double bonds and a second tail with only single bonds. The chain with double bond is said to be unsaturated, because it doesn’t contain the maximum number of hydrogen atoms. The fatty acid tail with only single bonds is said to be saturated. Since each double bond creates a small kink in the hydrocarbon tail, it makes it more difficult for the tails to pack against each other. Because of this, lipid bilayers that have unsaturated hydrocarbon tails are more fluid than bilayers that are saturated. Cholesterol molecules fill the spaces between phospholipid molecules which are left by unsaturated hydrocarbon tails; stiffening the bilayer and making it less permeable Asymmetry​: Asymmetry can occur on both sides of the lipid bilayer, when the inner and outer leaflet have different molecular architecture. Maintenance of proper lipid asymmetry is required for the mechanical stability of the membrane and for vesicular transport. Lipid rafts are specific examples. Phospholipids synthesized in ER are added to the cytosolic half of the bilayer. Half of the phospholipids are transferred to the opposite monolayer phosphatidcholyine, sphingomyelin, phosphatidylserine by flippases; creates asymmetry. Glycolipids are found only in the noncytosolic half of the bilayer. There they form part of a continuous protective coat of carbohydrate, glycocalyx. Glycolipids get their sugar group in the Golgi, and only lipids in the noncytosolic half of the bilayer get sugars added to them. Once a glycolipid molecule is made, it remains trapped in the monolayer without possibility of flippase; creates asymmetry. cholesterol also increases the asymmetry of lipid bilayer Glycocalyx or external coat or (glycolipids, glycoproteins, proteoglycans) The carbohydrates on the exterior surface of the cell the carbohydrate components of both glycoproteins and glycolipids are collectively referred to as the glycocalyx meaning “sugar coating”. Vishal Kumar Glycocalyx is a highly charged layer of membrane-bound biological macromolecules attached to a cell membrane. This layer functions as a barrier between a cell and its surrounding. Glycocalyx also serves as a mediator for cell-cell interactions and protects a cell membrane from the direct action of physical forces and stresses allowing the membrane to maintain its integrity. Glycocalyx is also involved in development and progression of many diseases. Glycocalyx is composed of glycosaminoglycans, proteoglycans and other glycoproteins bearing acidic oligosaccharides and terminal sialic acids. Most glycocalyx associated proteins are transmembrane that can be linked to the cytoskeleton. This linkage allows signal transduction from the external to the internal parts of a cell. Functions of membrane components: recognition and transport Most membrane functions are carried out by membrane proteins. There are different kinds of proteins. Transporter proteins ​transport nutrients, metabolites and ions across the bilayer. ​Anchor proteins ​anchor the membrane to macromolecules on both side. ​Receptor proteins ​detect chemical signals in the cell’s environment and relay them to the cell interior. ​Enzymes catalyze specific reactions, e.g. ATPase that catalyze the decomposition of ATP into ADP and a free phosphate ion. Membrane transport (simple diffusion, passive transport or facilitated diffusion, active transport: primary and secondary) Simple diffusion: ​In the process of ​diffusion​, a substance tends to move from an area of high concentration to an area of low concentration until its concentration becomes equal throughout a space. Over time the net movement of molecules will be out of the more concentrated area and into the less concentrated one, until the concentrations become equal. This process does not require any energy input; in fact, a concentration gradient itself is a form of stored potential energy, and this energy is used up as the concentrations equalize. Passive transport: ​Passive transport is a naturally occurring phenomenon and does not require the cell to exert any of its energy to accomplish the movement. In passive transport, substances move from an area of higher concentration to an area of lower concentration. The passive forms of transport, diffusion and osmosis, move materials of small molecular weight across membranes. Facilitated diffusion: ​Some molecules, such as carbon dioxide and oxygen, can diffuse across the plasma membrane directly, but others need help to cross its hydrophobic core. ( Due to integral protins ) In ​facilitated diffusion​, molecules diffuse across the plasma membrane with assistance from membrane proteins, such as channels and carriers. A concentration gradient exists for these molecules, so they have the potential to diffuse into or out of the cell by moving down it. However, because they are charged or polar, they Vishal Kumar can t cross the phospholipid part of the membrane without help. Facilitated transport proteins shield these molecules from the hydrophobic core of the membrane, providing a route by which they can cross. Two major classes of facilitated transport proteins are channels and carrier proteins. Channel proteins​ span the membrane and make hydrophilic tunnels across it, allowing their target molecules to pass through by diffusion. Channels are very selective and will accept only one type of molecule or a few closely related molecules for transport. Aquaporins​ are channel proteins that allow water to cross the membrane very quickly, and they play important roles in plant cells, red blood cells, and certain parts of the kidney where they minimize the amount of water lost as urine Another class of transmembrane proteins involved in facilitated transport consists of the carrier proteins. ​Carrier proteins​ can change their shape to move a target molecule from one side of the membrane to the other. Like channel proteins, carrier proteins are typically selective for one or a few substances. Active transport: primary One of the most important pumps in animals cells is the sodium-potassium pump Na​ ​-K​ ​ ATPase , which maintains the electrochemical gradient and the correct concentrations of Na​ ​ and K​ ​ in living cells. It utilizes energy in form of ATP to transport molecules across a membrane against their concentration gradient. The sodium-potassium pump moves two K​ ​ into the cell while moving three Na​ ​ out of the cell. Several things have happened as a result of this process. At this point, there are more sodium ions outside of the cell than inside and more potassium ions inside than out. This results in the interior being slightly more negative relative to the exterior. This difference in charge is important in creating the conditions necessary for the secondary process. The sodium-potassium pump is, therefore, an electrogenic pump a pump that creates a charge imbalance , creating an electrical imbalance across the membrane and contributing to the membrane potential. An important membrane adaption for active transport is the presence of specific carrier proteins or pumps to facilitate movement. There are three types of these proteins or transporters: ​uniporters, symporters, and antiporters​. A uniporter carries one specific ion or molecule. A symporter carries two different ions or molecules, both in the same direction. An antiporter also carries two different ions or molecules, but in different directions. All of these transporters can also transport small, uncharged organic molecules like glucose. These three types of carrier proteins are also found in facilitated diffusion, but they do not require ATP to work in that process. Active transport: Secondary Secondary active transport brings sodium ions, and possibly other compounds, into the cell. As sodium ion concentrations build outside the plasma membrane because of the action of Vishal Kumar the primary active transport process, an electrochemical gradient is created. If a channel protein exists and is open, the sodium ions will be pulled through the membrane. This movement is used to transport other substances that can attach themselves to the transport protein through the membrane. Many amino acids, as well as glucose, enter a cell this way. Structure and functions of cell nucleus​ ​ ​ ​ECB: , , , - OR​ ​ - , - , , , - ​ and ​ lecture of DGCI. week lecture nd​ rd​ supplements: nucleolus, nucleoskeleton nuclear transport processes see on the homepage Evolution of the nucleus. In bacteria, the singe DNA is typically attached to the plasma membrane. From the prokaryotic cell, the plasma membrane could have invaginated and formed a two-layered envelope surrounding the DNA. Envelope completely pinched of the membrane. The envelope also has channels called pores. Other portions of the same membrane formed the ER. Structure of nucleus Eukaryotic cells have a true nucleus, which means the cell’s DNA is surrounded by a membrane. Therefore, the nucleus houses the cell’s DNA and directs the synthesis of proteins and ribosomes. The nuclear envelope is a double-membrane structure that constitutes the outermost portion of the nucleus. Both the inner and outer ( doble layer membranes of the nuclear envelope are phospholipid bilayers. The nuclear envelope is punctuated with pores that control the passage of ions, molecules, and RNA between the ) I 4 phospholipids nucleoplasm and cytoplasm. The nucleoplasm is the semi-solid fluid inside the nucleus where we find the chromatin and the nucleolus. Furthermore, chromosomes are structures within the nucleus that are made up of DNA, the genetic material. In prokaryotes, DNA is organized into a single circular chromosome. In eukaryotes, chromosomes are linear structures. Vishal Kumar Nuclear envelope The nuclear envelope is a highly regulated membrane barrier that separates the nucleus from the cytoplasm. It consists of double layer inner and outer membrane of a bilayer of phospholipids and many proteins. Some of these proteins are known as Nups nucleoporins , where exchange of proteins as RNA and ribonucleoprotein between the nucleus and cytoplasm occur. The inner surface of the nuclear envelope has a protein lining called the nuclear lamina. The nuclear envelope contains many nuclear pores, which form transport routes which are fully permeable for smaller molecules up to the size of small proteins. However, they form a selective barrier against larger molecules. Each pore is surrounded by a cooperating protein structure called the nuclear pore complex, which does the selection of which proteins may enter the nucleus. Typical molecules that pass the pores are nucleotides DNA building blocks, ATP and histones. During mitosis, the nuclear envelope must be completely degenerated to allow the mitotic spindle to attach to the chromosomes, and then be rebuilt. Nuclear lamina The nuclear lamina is a dense network of fibres on the inner side of the inner nucleic membrane. Thus, it has an important structural role and is also known as the nuclear skeleton. The lamina has anastomosis with the NE through proteins especially with the inner NE. Besides providing mechanical support, the nuclear lamina regulates events such as DNA replication and cell division. It also participates in chromatin organization and it anchors the nuclear pore complexes on the NE. Various functions have been suggested for the nuclear lamina and intranulear filaments:. maintenance of nuclear envelope and nuclear shape;. spatial organisation of nuclear pores within the nuclear membrane;. regulation of transcription;. anchoring of interphase heterochromatin;. role in DNA replication; Interestingly, lamins are found within the nucleoplasm intranuclear filaments and these lamin foci are often associated with sites of DNA replication. Nuclear pores The nuclear envelope in all eukaryotic cells is perforated by nuclear pores that form the gates through which all molecules enter or leave the nucleus. A nuclear pore is a large, elaborate structure composed of different proteins. Each pore contains water-filled passages through which small water-soluble molecules can pass freely. Larger molecules, such as RNA and proteins, must display an appropriate sorting signal to pass through the pore. The signal sequence that directs a protein from the cytosol into the nucleus, is called a nuclear localization signal. Vishal Kumar Eu- and heterochromatin, chromosome territories, further chromatin organization (chromatin fiber, loops, rosette) Basic ✓ Chromatin is composed of DNA and histones that are packaged into thin, stringy fibers. These chromatin fibers are not condensed but can exist in either a compact form heterochromatin or less compact form euchromatin. Processes including DNA replication, transcription, and recombination occur in euchromatin. During cell division, chromatin condenses to form chromosomes. Histones help organize DNA into structures called nucleosomes by providing a base on which the DNA can be wrapped around. The nucleosome is further folded to produce a chromatin fiber. Chromatin fibers are coiled and condensed to form chromosomes. Chromatin makes it possible for many cell processes to occur including DNA replication, cell division etc. The major difference between heterochromatin and euchromatin is that heterochromatin is such part of the chromosomes, which is a firmly packed form and are genetically inactive, while euchromatin is an uncoiled loosely packed form of chromatin and are genetically active. Constitutive heterochromatin is the stable form of heterochromatin, i.e. it does not loosen up to form euchromatin, and contains repeated sequences of DNA called satellite DNA. It can be found in centromeres and telomeres. Facultative heterochromatin, on the other hand, is reversible, i.e. its structure can change depending on the cell cycle. Nucleosome structure Proteins that bind to DNA to form chromosomes are divided into histones and non-histones. Histones are responsible for the first level of chromatin packing the packing from DNA strand to nucleosomes. Nucleosomes are small packs of proteins histones packed with DNA around like loops and between them. Between the nucleosomes are linker-DNA. Each nucleosome core particle consists of a complex of eight different histone proteins. Two molecules each of H A, H B, H and H , and the double stranded DNA around the core particle. The histones are positive charged, giving them the ability to bind tightly to the negative charged backbones of the DNA. But the DNA is packed even tighter, by a fifth histone H , which pull several nucleosome strands on top of each other. Histones contain a special amino acid sequence on their surface which allow them entry to the cell nucleus through the NE. ​Ill stration ello are histones red is DNA Vishal Kumar Nucleoskeleton: nuclear lamina, connection to nuclear membrane and intranuclear lamins. The nucleoskeleton of an animal cell is made from intermediate filaments. It contains nuclear lamina and intranuclear filaments. The ​nuclear lamina is a scaffold-like network of protein filaments​ surrounding the nuclear periphery beneath the inner nuclear membrane. This scaffold is made of mostly the ancient intermediate filament proteins, ​lamin A/C and B​, which together form a complex meshwork. Lamin A and C has a common gene and the different protein coding mRNAs are made by alternative splicing. Nuclear lamin B has a receptor in the inner nuclear membrane lamin B receptor , so lamina layer makes connection to the membrane and to the chromatin as well perinuclear heterochromatin. During cell cycle in prophase of mitosis lamins are phosphorylated and nuclear envelope is broken. In telophase lamins are dephosphorylated and nuclear envelope is reorganised. These intermeidier filament forming lamins are found inside the nucleus as well, making a loose network of filaments called intranuclear filaments. Various functions have been suggested for the nuclear lamina and intranulear filaments:. maintenance of nuclear envelope and nuclear shape;. spatial organisation of nuclear pores within the nuclear membrane;. regulation of transcription;. anchoring of interphase heterochromatin;. role in DNA replication; Interestingly, lamins are found within the nucleoplasm intranuclear filaments and these lamin foci are often associated with sites of DNA replication. Autosomal dominant inherited form of ​Emery-Dreifuss muscular dystrophy​ EDMD have been linked to mutations in lamin A/C. Another type of disease, called ​Hutchinson-Guilford progeria​ is caused by lamin A mutation, too. Mechanism and regulation of nuclear (gated) transport: nuclear export and import signals, kariopherins (importin, exportin), role of Ran-GTP and Ran-GDP Nuclear envelope of all eucaryotes is perforated by large structures known as ​nuclear pore complexes​. It is thought to be composed of about different proteins in many copies. These proteins are called ​nucleoporins,​ that are arranged in an octagonal symmetry. Each pore complex contains many aqueous channels through which small water-soluble molecules can passively diffuse. Small molecules daltons or less diffuse so fast that the nuclear envelope can be considered to be freely permeable to them. Vishal Kumar The mechanism of macromolecular transport across nuclear pore complexes is fundamentally different from the transport mechanisms of other proteins across other membranes of e.g. ER, mitochondrion. It occurs through large aqueous pore rather than through a protein transporter spanning one or more lipid bilayers. For this reason, ​nuclear transport is a gated transport, ​where nuclear ​proteins are carried ​through the pore complex being a fully folded conformation​. Some contributors of the transport are; cargo, karyopherins, nucleoporins and ran proteins. Cargo is the transported molecule. It comes with a signal. Either NLS nuclear localization signal for import , NES nuclear export signal for export. Sequences of amino acids work as the signal. Karyopherins are the transporters in form of importin and exportin, which binds to the cargo and transports it in or out. The nucleoporins are part of the NE and serve as a way in or out of the nucleus. Ran proteins small or monomeric G proteins , regulate the transport processes G protein can bind GTP or GDP. The ran proteins are the drivers powerhouse of the transport. Ran GDP moves from the cytoplasm into the nucleus without binding to the importin cargo ( molecule to be transported) complex. It creates a Ran GDP gradient wich helps the transport. ​In the nucleus Ran exchanges its GDP to GTP by the help of Ran GEF that causes the release of cargo from importin receptor. Ran-GTP itself is bound to the exportin cargo complex in the nucleus and exported to the cytosol.​ There Ran-GAP converts Ran-GTP to Ran-GDP and exportin releases both its cargo and Ran-GDP. Then Ran-GDP is able to move into the nucleus again. ​So inaequal distribution of Ran GDT/GTP is the regulator for export and import. Role of Ran proteins: ​The gated transport through the nuclear pores is driven by so called Ran proteins that are members of a protein family called ​monomeric G proteins​. ​G proteins exist in two conformational states,​ depending on whether ​GDP ​or​ GTP ​is bound to them. Conversion between the two states is triggered by two Ran specific regulatory proteins: a cytosolic ​GTPase acti ating protein ​Ran GAP​ ​ that triggers GTP hydrolysis and thus converts Ran-GTP to Ran-GDP, and a nuclear ​g anine e change factor ​Ran GEF​ ​ that promotes the exchange of GDP for GTP and thus converts Ran-GDP to Ran-GTP. Ran itself is found both in cytosol and nucleus, but in different forms. ​Since ​Ran GAP​ is located in the cytosol and ​Ran GEF​ is located in the nucleus, the cytosol primarily contains Ran-GDP, and the nucleus primarily contains Ran-GTP which maintains a Ran-GDP and Ran-GTP gradient between the compartments. Vishal Kumar Structure and function of nucleolus. (FC, DFK, DGK, rRNA synthesis and processing). The nucleolus is the most obvious structure in the nucleus of a eucaryotic cell viewed in the light microscope. It is one of the biggest subcompartment inside the nucleus one of the nuclear bodies in the interchromatin area. Nucleolus is the site of rRNA synthesis and processing and their assembly with ribosomal proteins which process is called ribosome biogenesis. Unlike other organelles in the cell, it is not covered by a membrane instead, it is a large aggregation of macromolecules, including rRNA genes, precursor rRNAs, mature rRNAs, rRNA-processing enzymes, snoRNPs, ribosomal protein and partly assembled ribosomal subunits. The close association of all these components presumably allows the assembly of ribosomes to occur rapidly and smoothly. Parts of nucleolus (visible under electronmicroscope):. fibrillar center (FC) /pars amorpha /NOR/: place of transcription for the rRNA genes by RNA polymerase I and the transcription factors transcriptosome I. Marker protein of this part is UBF transcription factor.. dense fibrillar component (DFC)/ pars fibrosa/: splicing and modification of pre-rRNA made by the snoRNPs small nucleolar ribonucleoproteins. Marker protein of this part is fibrillarin.. granular component (GC)/ pars granulosa/: matured rRNAs and ribosomal proteins are assembled into ribosomal subunits. Marker protein of this part is B wich is a transporter-molecule. Then the ribosomal subunits are exported from the nucleus to the cytosol.. Nucleolus associated heterochromatin or chromatin /pars chromosoma/​: most of this heterochromatin has no functional relation to the nucleolus this part contains functionally different chromatin of , , , and human chromosomal DNA. Functions of the nucleus:. Transcription of pre-rRNA. : modification of pre-rRNA. : Assembly with proteins. : Transport Characteristics of eukaryotic transcription (eukaryotic RNA polymerases, mRNA processing: cap and polyA formation, steps of splicing, splicosome) RNA: ​The process by which DNA is copied to RNA is called transcription. RNA is a linear polymer made of four different types of nucleotide subunits linked together by phosphodiester bonds. The difference between RNA and DNA is that RNA contains the sugar ribose; ribonucleic acid. RNA has the bases A, C, G and U. DNA is double-stranded, while RNA is single-stranded. Transcription: ​All the RNA in a cell is made by transcription. Transcription begins with the opening and unwinding of a small portion of the DNA double helix to expose the bases on each DNA strand. One of the DNA strands acts as a template for RNA synthesis. Ribonucleotides are added to the growing RNA chain; the nucleotide sequence of the RNA Vishal Kumar chain is determined by the complementary base-pairing of the DNA template. The RNA chain produced by transcription has a nucleotide sequence exactly complementary to the strand of DNA used as the template. Polymerase: ​The enzymes that carry out transcription are called RNA polymerases. RNA polymerases catalyze the formation of the phosphodiester bonds that link the nucleotides together and form the sugar-phosphate backbone of the RNA chain. The difference between DNA polymerase and RNA polymerase, is that the RNA polymerase can start an RNA chain without a primer. This difference may exist because transcription does not need to be as accurate as DNA replication; RNA is not used as permanent storage of genetic information in cells. RNAs: ​Most of the genes carried in a cell’s DNA specify the amino acid sequence of proteins, and the RNA molecules that are copied from these genes are called messenger RNA. Each mRNA usually carries information transcribed from just one gene, coding for a sing protein. Ribosomal RNA rRNA forms the core of the ribosomes, on which mRNA is translated into protein, and transfer RNA tRNA forms the adaptors that select amino acids and hold them in place for a ribosome for incorporation into protein. mRNA processing: ​In bacteria, RNA transcripts are ready to act as messenger RNAs and get translated into proteins right away. In eukaryotes, things are a little more complex. The molecule that s directly made by transcription in one of your eukaryotic cells is called a ​pre-mRNA​, reflecting that it needs to go through a few more steps to become an actual messenger RNA mRNA. These are: Both ends of a pre-mRNA are modified by the addition of chemical groups. The group at the beginning end is called a cap, while the group at the end end is called a tail. Both the cap and the tail protect the transcript and help it get exported from the nucleus and translated on the ribosomes protein-making machines found in the cytosol. The ​ ’ cap​ is added to the first nucleotide in the transcript during transcription. The cap is a modified guanine G nucleotide, and it protects the transcript from being broken down. It also helps the ribosome attach to the mRNA and start reading it to make a protein. The end of the RNA forms in kind of a bizarre way. When a sequence called a ​polyadenylation signal​ shows up in an RNA molecule during transcription, an enzyme chops the RNA in two at that site. Another enzyme adds about - adenine A nucleotides to the cut end, forming a ​poly-A tail​. The tail makes the transcript more stable and helps it get exported from the nucleus to the cytosol. The third big RNA processing event that happens in your cells is ​RNA splicing​. In RNA splicing, specific parts of the pre-mRNA, called ​introns​ are recognized and removed by a protein-and-RNA complex called the ​spliceosome​. Introns can be viewed as junk sequences that must be cut out so the good parts version of the RNA molecule can be assembled. The pieces of the RNA that are not chopped out are called ​exons​. The exons are pasted together by the spliceosome to make the final, mature mRNA that is shipped out of the nucleus. Vishal Kumar A key point here is that it s ​onl ​ the exons of a gene that encode a protein. The introns actually ​ha e​ to be removed in order for the mRNA to encode a protein with the right sequence. If the spliceosome fails to remove an intron, an mRNA with extra junk in it will be made, and a wrong protein will get produced during translation. The splicing, cutting and pasting is performed by the spliceosome; enzyme made of protein and RNAs that carries out RNA splicing in the cell. Small nuclear RNAs snRNAs are packaged with additional proteins to form small nuclear ribonucleoprotein particles snRNPS ‘’snurps’’. The snRNPs form the core of the spliceosome. Once it s completed these steps, the RNA is a mature mRNA. It can travel out of the nucleus and be used to make a protein. Structure of eukaryotic mRNA. Structure and functions of endoplasmic reticulum (ER)​ ​ ​ ​ECB​: , , OR ​ - , – 0 , 0 - 0 ​.​ ​ lecture of DGCI th​ ​ week practice) th​ The endoplasmic reticulum ER is the major site of synthesis of new membranes in the cell. Large areas of the ER have ribosomes attached to the cytosolic surface and are known as rough ER. The ribosomes are actively synthesizing proteins that are delivered into the ER lumen or membrane. The smooth ER lacks ribosomes. It is the site of steroid hormone synthesis in cells of the adrenal gland and the site of organic molecules, as alcohol, are detoxified in liver cells. In many eukaryotic cells, the smooth ER also regulates the Ca​ ​ion concentration in the cytoplasm. Vishal Kumar Structure and function of the smooth ER (sER: phospholipid synthesis, Ca storage, detoxification, steroid synthesis, glucose metabolism) The smooth endoplasmic reticulum SER is continuous with the RER but has few or no ribosomes on its cytoplasmic surface. Functions of the SER include synthesis of carbohydrates, lipids phospholipids, cholesterol , and steroid hormones; detoxification of medications and poisons liver ; and storage of calcium ions. In muscle cells, a specialized SER called the sarcoplasmic reticulum is responsible for storage of the calcium ions that are needed to trigger the coordinated contractions of the muscle cells.​ ​The smooth ​endoplasmic reticulum​ also contains the enzyme ​glucose​- -phosphatase, which converts ​glucose​- -phosphate to ​glucose​, a step-in gluconeogenesis liver cells. Transport of membrane lipids The ​ER​ membrane synthesizes nearly all of the major classes of lipids, including both phospholipids and cholesterol, required for the production of new cell membranes. Although mitochondria modify some of the lipids they import, they do not synthesize lipids from scratch; instead, their lipids have to be imported from the ER, either directly, or indirectly by way of other cell membranes. In either case, special mechanisms are required for the transfer. Water-soluble carrier proteins called phospholipid exchange proteins or ​phospholipid transfer proteins ​ transfer individual phospholipid molecules between membranes. Each exchange protein recognizes only specific types of phospholipids. It functions by “extracting” a molecule of the appropriate phospholipid from a membrane and diffusing away with the lipid buried within its lipid-binding site. When it encounters another membrane, the exchange protein tends to discharge the bound phospholipid molecule into the new lipid bilayer. The ER also produces cholesterol and ceramide. ​Ceramide​ is made by condensing the amino acid serine with a fatty acid to form the amino alcohol ​sphingosine ​ a second fatty acid is then added to form ceramide. The ceramide is exported to the Golgi apparatus, where it serves as a precursor for the synthesis of two types of lipids: oligosaccharide chains are added to form ​gl cosphingo lipids​ glycolipids , and phosphocholine head groups are transferred from phosphatidylcholine to other ceramide molecules to form ​sphingom elin​. Thus, both Vishal Kumar glycolipids and sphingomyelin are produced relatively late in the process of membrane synthesis. Because they are produced by enzymes exposed to the Golgi lumen and are not substrates for lipid translocators, they are found exclusively in the noncytosolic leaflet of the lipid bilayers that contain them. Comparison of eu- and prokaryotic ribosomes Prokaryotic ribosomes are smaller s than eukaryotic ribosomes s Prokaryotic ribosomes consist of S and S, whereas eukaryotic ribosomes consist of S and S. In eukaryotic cells, ribosomes are in free and bound form, while prokaryotes only have the free form. Protein sorting, targeting; role of signals The rough ER is a sort of mail room for proteins that get sent to the ER, they are sorted according to their destinations, then forwarded to the appropriate cellular compartment for some proteins, further sorting occurs in the Golgi apparatus, as well. Proteins that are to be ​secreted​ out of the cell, as well as proteins bound for the plasma ​membrane​, the ​Golgi​ apparatus and ​l sosomes​ plus those destined for the ​ER​ itself are sent to the ER. Those that are travelling beyond the ER move from the ER to the Golgi, and from there to the outside of the cell in secretory vesicles, if they are secreted proteins. If they are intended for the lysosome, the vesicles fuse with the lysosome to deliver them. Structure and function of the rough ER (rER) The ​rough endoplasmic reticulum​ ​rough ER​ gets its name from the bumpy ribosomes attached to its cytoplasmic surface. Ribosomes transfer their newly synthesized proteins into the lumen of the RER where they undergo structural modifications, such as folding or the acquisition of side chains. These modified proteins will be incorporated into cellular membranes the membrane of the ER or those of other organelles or secreted from the cell such as protein hormones, enzymes. The RER also makes phospholipids for cellular membranes. If the phospholipids or modified proteins are not destined to stay in the RER, they will reach their destinations via transport vesicles that bud from the RER’s membrane. Since the RER is engaged in modifying proteins such as enzymes, for example that will be Vishal Kumar secreted from the cell, the RER is abundant in cells that secrete proteins. This is the case with cells of the liver, for example. Functions of rough ER: - protein synthesis - co-translational translocation - integration of transmembrane protein - post-translational protein modification.g. proteolytic cleavage, N-glycosilation - protein folding chaperons - quality control - transportation of proteins Protein synthesis in ER bounded ribosome mRNA transcribed from the nucleus gets exported into the cytosol through a nuclear pore. When it comes into contact with a ribosome and binds to it, the process of translation can begin. Many ribosomes bind into the mRNA, forming a polyribosome. There are two populations of polyribosome that share the same pool of ribosomal subuntis, the free ribosomes and the membrane-bound ribosomes. Free ribosomes translate mRNA into proteins that are taken up by the cytoplasm or other organelles for the functioning of the cell. Membrane-bound ribosomes synthesize proteins that are translocated into the lumen of the ER. Proteins destined for the Golgi, endosomes, lysosomes and the cell surface, all first enter the ER from the cytosol. Two kinds of proteins are transferred from the cytosol to the ER; water-soluble proteins and prospective transmembrane proteins. Water-soluble proteins are completely translocated across the ER membrane and are released into the ER lumen. They are destined either for secretion or for the lumen of an organelle. The transmembrane proteins are only partly translocated across the ER membrane and become embedded in it. They are destined to reside in either the ER membrane, membrane of another organelle or plasma membrane. All these proteins are initially directed to the ER by an ER signal sequence. Function of SRP and SRP receptor The ER signal sequence is guided to the ER membrane with the aid of at least two protein components; : a signal-recognition particle SRP , present in the cytosol, which binds to the ER signal sequence when it is exposed on the ribosome, and : an SRP receptor, embedded in the membrane of the ER, which recognizes the SRP. Binding of an SRP to a signal sequence causes protein synthesis by the ribosome to slow down, until the ribosome and its bound SRP locate an SRP receptor on the ER. After the binding to its receptor, the SRP is released and protein synthesis recommences, with the polypeptide now being threaded into the Vishal Kumar lumen of ER through a translocation channel in the ER membrane. Thus, the SRP and SRP receptor function as molecular matchmakers, connecting ribosomes that are synthesizing proteins containing ER signal sequences to available ER translocation channels. Cotranslational transmembrane transport Processing of proteins in the lumen of rER: N-glycosylation; activity of chaperons in folding, quality control Protein folding in the ER lumen is initiated by the addition of an oligosaccharide chain to the polypeptide and has assisted by a folding chaperone called calnexin. Many of the proteins that enter the ER lumen or membrane are converted to glycoproteins in the ER by the attachment of short oligosaccharide side chains. This process of glycosylation is carried out by glycosylating enzymes present in the ER but not in the cytosol. Oligosaccharides can protect the protein from degradation, hold it in the ER until it is folded or guide it to the appropriate organelle. An oligosaccharide branch containing a total of sugars is attached to all proteins that carry the appropriate site for glycosylation. Most proteins that enter the ER are destined for other locations; they are packaged into transport vesicles that bud from the ER and fuse with the Golgi apparatus. Exit from the ER is highly selective. Proteins that fold incorrectly are retained in the ER by binding to chaperone proteins. Interaction with chaperones hold the proteins in the ER until proper folding occurs; if this does not happen, proteins are eventually degraded. In this way, the ER controls the quality of the proteins that it exports to Golgi. Vishal Kumar Structure and function of proteasomes. Misfolded proteins are transported to the cytosol where they are degraded by the multienzyme complex; proteasome. Proteasomes are protein complexes which degrade unneeded or damaged proteins by proteolysis; a chemical reaction that breaks peptide bonds.. Structure and function of Golgi.​ ​ ​ECB: ​ - OR ​ 0– ​.​ ​th​ lecture of DGCI th ​ week practice​ Polarity of Golgi apparatus The Golgi apparatus is very polar. Membranes at one end of the stack differ in both composition and in thickness from those at the other end. One end cis face acts as the receiving department while the other trans face acts as the shipping department. The cis face is closely associated with the ER. Golgi subcompartmets (CGN, cis, medial, trans Golgi, TGN) Vishal Kumar The Golgi apparatus is usually located near the cell nucleus and plasma membrane. It consists of a collection of flattened, membrane-enclosed sacs cisternae. The entry of the golgi, cis, is near the ER. The first outermost cisterna is the; cis Golgi network. Then comes the cisternae; first cis-cisterna, medial-cisterna and then trans-cisternae. After that comes the trans Golgi network. The exit of the Golgi, trans, is pointed towards the plasma membrane. Golgi functions: transport, sorting. M- phosphorilation, O-glycosylation, modification of N- oligosaccharide chains anf lipids.) The proteins and lipids received at the cis face arrive in clusters of fused vesicles. These fused vesicles migrate along microtubules through a special trafficking compartment, called the vesicular-tubular cluster, that lies between the endoplasmic reticulum and the Golgi apparatus. When a vesicle cluster fuses with the cis membrane, the contents are delivered into the lumen of the cis face cisterna. As proteins and lipids progress from the cis face to the trans face, they are modified into functional molecules and are marked for delivery to specific intracellular or extracellular locations. Some modifications involve cleavage of oligosaccharide side chains followed by attachment of different sugar moieties in place of the side chain. Other modifications may involve the addition of fatty acids or phosphate groups phosphorylation or the removal of monosaccharides. The different enzyme-driven modification reactions are specific to the compartments of the Golgi apparatus. In the final stage of transport through the Golgi apparatus, modified proteins and lipids are sorted in the trans Golgi network and are packaged into vesicles at the trans face. These vesicles then deliver the molecules to their target destinations, such as lysosomes or the cell membrane. Some molecules, including certain soluble proteins and Vishal Kumar secretory proteins, are carried in vesicles to the cell membrane for exocytosis release into the extracellular environment. Secretion (constitutive and regulated secretion, exocytosis) In all eukaryotic cells, a steady stream of vesicles buds from the trans-Golgi-network and fuses with the plasma membrane. This ​constitutive exocytosis pathway ​operates continually and supplies newly made lipids and proteins to the plasma membrane. The constitutive pathway also carries proteins to the cell surface to be released to the outside; a process called secretion. While the constitutive exocytosis pathway, which operates continually in all eukaryotic cells, there is also a regulated exocytosis pathway, ​which operates only in cells that are specialized for secretion.. Vesicular transport​ ​ ​ ​ECB​: - ​ R​ 0 - 0 ​. ​ ​O ​ lecture of DGCI​ th​ Most important molecules in the mechanism and the regulation of vesicular transport: signals, receptors, coat proteins, cytoskeletal components, SNARE-s, Rab proteins, NSF. Transport from the ER to the Golgi and from the Golgi to other compartments of the endomembrane system is carried out by continual budding and fusion of transport vesicles. Vesicular transport between endomembrane system compartments is highly organized. A major outward secretory pathway starts with the synthesis of proteins on the ER membrane and their entry into the ER, and it leads through the Golgi to the cell surface; at the Golgi, a Vishal Kumar side branch leads off through endosomes to lysosomes. A major inward endocytic pathway, which is responsible for the ingestion and degradation of extracellular molecules, moves materials from the plasma membrane, through endosomes to lysosomes. Vesicles that bud from membranes usually have a distinctive protein coat on their cytosolic surface and are therefore called coated vesicles. The best-studied vesicles are those that have coats made largely of protein clathrin. These clathrin-coated vesicles bud from the Golgi on the outward secretory pathway and from the plasma membrane on the inward endocytic pathway. Proteins called adaptins secure the clathrin coat to vesicles and help select cargo molecules for transport. Molecules for onward transport carry specific transport signals that are recognized by cargo receptors in the compartment membrane. Adaptin help capture cargo moleculues by trapping the cargo receptors that bind them. After transport vesicles buds from the membrane, it must find its way to the correct destination and deliver the contents. Once a vesicle has reached its target, it must recognize and dock with the organelle, so it can fuse the membranes and unload the cargo. Transport vesicles have ​Rab proteins ​on their surfaces which have to be recognized by ​tethering proteins ​on the surface of the target membrane. This ensures that vesicles only fuse with the correct membrane. There also ​SNAREs ​proteins which are transmembrane proteins. Once the tethering protein captures a vesicle by Rab proteins, the SNAREs on the vesicles interact with the SNAREs on the target membrane. Once a transport vesicle has recognized its target membrane and docked there, the vesicle has to fuse with the membrane and deliver its cargo. Vishal Kumar. Endocytosis. Intracellular digestion​ ​ ​ ​ECB​: - OR ​ - 0​. ​ ​ lecture of DGCI​ ​ th th​ ​ week practice) Types of endocytosis: pinocytosis, and phagocytosis Endocytosis ​endo​ internal, ​c tosis​ transport mechanism is a general term for the various types of active transport that move particles into a cell by enclosing them in a vesicle made out of plasma membrane. There are variations of endocytosis, but all follow the same basic process. First, the plasma membrane of the cell invaginates folds inward , forming a pocket around the target particle or particles. The pocket then pinches off with the help of specialized proteins, leaving the particle trapped in a newly created vesicle or vacuole inside the cell. Phagocytosis cellular eating involves the ingestion of large particles via large vesicles called phagosomes. The phagosomes fuse with lysosomes, where the food particles are digested and broken down into its basic components, which can be used by the cell. Pinocytosis literally, “cell drinking” is a form of endocytosis in which a cell takes in small amounts of extracellular fluid. Pinocytosis occurs in many cell types and takes place continuously, with the cell sampling and re-sampling the surrounding fluid to get whatever nutrients and other molecules happen to be present. Pinocytosed material is held in small vesicles, much smaller than the large food vacuole produced by phagocytosis. Receptor mediated endocytosis Receptor-mediated endocytosis​ is a form of endocytosis in which receptor proteins on the cell surface are used to capture a specific target molecule. The receptors, which are transmembrane proteins, cluster in regions of the plasma membrane known as coated pits. This name comes from a layer of proteins, called coat proteins, that are found on the cytoplasmic side of the pit. Clathrin is the best-studied coat protein. When the receptors bind to their specific target molecule, endocytosis is triggered, and the receptors and their attached molecules are taken into the cell in a vesicle. The coat proteins participate in this process by giving the vesicle its rounded shape and helping it bud off from the membrane. Receptor-mediated endocytosis allows cells to take up large amounts of molecules that are relatively rare present in low concentrations in the extracellular fluid. Although receptor-mediated endocytosis is intended to bring useful substances into the cell, other, less friendly particles may gain entry by the same route. Flu viruses, diphtheria, and cholera toxin all use receptor-mediated endocytosis pathways to gain entry into cells. Vishal Kumar Digestion of selfcomponents: autophagocytosis Autophagy is a normal physiological process in the body that deals with destruction of cells in the body. It maintains homeostasis or normal functioning by protein degradation and turnover of the destroyed cell organelles for new cell formation. Delivers the cytoplasmic constituents to lysosome. Difference between autophagocytosis and autolysis Autophagy usually refers to an ordered and purposeful digestion of cellular components. It s basically the way a cell can deal with unused or poorly folded proteins. This is a normal cellular process. Autolysis on the other hand occurs when digestive enzymes leak out of lysosomes and start destroying the cell or tissues. Functions of the endosomal – lysosomal compartment (early and late endosome, lysosome): sorting, digestion The endosomal compartment acts as the main sorting station in the inward endocytic pathway, just as the trans Golgi network serves this function in the outward secretory pathway. The acidic environment of the endosome plays a crucial part in the sorting process by causing many but not Vishal Kumar all receptors to release their bound cargo. The routes taken by receptors once they have entered an endosome differ according to the type of receptor: most are returned to the same plasma membrane domain from which they came, as is the case for the LDL receptor discussed earlier; some travel to lysosomes, where they are degraded; and some proceed to a different domain of the plasma membrane, thereby transferring their bound cargo molecules across the cell from one extracellular space to another, a process called transcytosis. Cargo proteins that remain bound to their receptors share the fate of their receptors. Cargo that dissociates from receptors in the endosome is doomed to destruction in lysosomes, along with most of the contents of the endosome lumen. Late endosomes contain some lysosomal enzymes, so digestion of cargo proteins and other macromolecules begins in the endosome and continues as the endosome gradually matures into a lysosome: once it has digested most of its ingested contents, the endosome takes on the dense, rounded appearance characteristic of a mature, “classical” lysosome. PRACTICAL: pH for early endosome and pH for late endosomes. The use of pH indicator dyes e.g. Congo Red can distinguish early and late endosomes, because of PH! Transcytosis Transcytosis​ is a type of transcellular transport in which various macromolecules are transported across the interior of a cell. Macromolecules are captured in vesicles on one side of the cell, drawn across the cell, and ejected on the other side.. Mitochondria and peroxisomes​ ​ ​ ​ECB​:​ ​ - , - , - , OR ​ - , – ​ molecular details are not needed. ​ ​ lecture of DGCI​ th​ Structure and function of mitochondria An individual mitochondrion is bounded by two highly specialized membranes. These membranes, called the outer and inner mitochondrial membranes, create two mitochondrial compartments: a large internal space called the Vishal Kumar matrix and a much narrower intermembrane space. The outer membrane contains many molecules of a transport protein called porin. As a result, the outer membrane is like a sieve that is permeable to all molecules of daltons or less, including small proteins. This makes the intermembrane space chemically equivalent to the cytosol with respect to the small molecules and inorganic ions it contains. In contrast, the inner membrane, like other membranes in the cell, is impermeable to the passage of ions and most small molecules, except where a path is provided by specific membrane transport proteins. The mitochondrial matrix therefore contains only molecules that are selectively transported into the matrix across the inner membrane, and so its contents are highly specialized. The inner mitochondrial membrane is the site of oxidative phosphorylation, and it contains the proteins of the electron-transport chain, the proton pumps, and the ATP synthase required for ATP production. It also contains a variety of transport proteins that allow the entry of selected small molecules such as pyruvate and fatty acids that will be oxidized by the mitochondrion into the matrix. The inner membrane is highly convoluted, forming a series of infoldings known as cristae that project into the matrix space. These folds greatly increase the surface area of the membrane. Functions: ▪ ATP synthesis ▪ Regulation of Ca​ ​ levels in the cell (cation granula) ▪ Lipid homeostasis (lipid oxidation, steroid synthesis) ▪ Nucleotide metabolism ▪ Amino acid metabolism ▪ FE-S synthesis (Hem) ▪ Ubiquinone synthesis ▪ Cofactor synthesis ▪ Apoptosis ▪ Aging ▪ Heat production Vishal Kumar Special molecules of mitochondrial subcompartments: eg.: porin, cardiolipin Porin is a protein which is present in the outer membrane of the mitochondria. It forms wide aqueous channels through the lipid bilayer. As a result, the outer membrane is permeable to all molecules of daltons or less including small proteins. This makes the intermembrane space equivalent to the cytosol. Cardiolipin is a unique phospholipid which is localized at level of the inner mitochondrial membrane where it is biosynthesized. This phospholipid is associated with membranes designed to generate an electrochemical gradient that is used to produce ATP. Such membranes include the bacterial plasma membrane and inner mitochondrial membrane.. Cardiolipin has been shown to interact with a number of inner mitochondrial membrane proteins including the respiratory chain complexes and substrate carriers. Main biochemical processes (not in details) of mitochondria: citric acid cycle, oxidative phosphorylation, chemiosmotic coupling, respiratory chain, ATP synthesis Mitochondria produce ATP through process of cellular respiration specifically, aerobic respiration, which requires oxygen. The citric acid cycle, or Krebs cycle, takes place in the mitochondria. This cycle involves the oxidation of pyruvate, which comes from glucose, to form the molecule acetyl-CoA. Acetyl-CoA is in turn oxidized and ATP is produced. The citric acid cycle reduces nicotinamide adenine dinucleotide NAD​ ​ to NADH. NADH is then used in the process of oxidative phosphorylation, which also takes place in the mitochondria. Electrons from NADH travel through protein complexes that are embedded in the inner membrane of the mitochondria. This set of proteins is called an electron transport chain. Energy from the electron transport chain is then used to transport proteins back across the membrane, which power ATP synthase to form ATP. The amount of mitochondria in a cell depends on how much energy that cell needs to produce. Muscle cells, for example, have many mitochondria because they need to produce energy to move the body. Red blood cells, which carry oxygen to other cells, have none; they do not need to produce energy. Mitochondria are analogous to a furnace or a powerhouse in the cell because, like furnaces and powerhouses, mitochondria produce energy from basic components in this case, molecules that have been broken down so that they can be used. Mitochondria have many other functions as well. They can store calcium, which maintains homeostasis of calcium levels in the cell. They also regulate the cell’s metabolism and have roles in apoptosis controlled cell death , cell signaling, and thermogenesis heat production Chemiosmosis is described as one of the mechanisms by which ATP is produced. As the electrons pass through the electron transport chain, energy is released, which is used to establish a proton gradient across a selectively-permeable membrane. The proton gradient drives the protons hydrogen ions to move down the gradient, releasing the energy that is in turn captured in the terminal phosphate bonds of ATP. Vishal Kumar Protein (posttranslational transmembrane transport) and lipid transport into mitochondria Although both organelles contain their own genomes and make some of their own proteins, most mitochondrial and chloroplast proteins are encoded by genes in the nucleus and are imported from the cytosol. These proteins usually have a signal sequence at their N-terminus that allows them to enter their specific organelle. Proteins destined for either organelle are translocated simultaneously across both the inner and outer membranes at specialized sites where the two membranes contact each other. Each protein is unfolded as it is transported, and its signal sequence is removed after translocation is complete Chaperone proteins inside the organelles help to pull the protein across the two membranes and to fold it once it is inside. The insertion of transmembrane proteins into the inner membrane, for example, is guided by signal sequences in the protein that start and stop the transfer process across the membrane, as we describe later for the insertion of transmembrane proteins in the ER membrane. The growth and maintenance of mitochondria and chloroplasts require not only the import of new proteins but also the incorporation of new lipids into the organelle membranes. Most of their membrane phospholipids are thought to be imported from the ER, which is the main site of lipid synthesis in the cell. Phospholipids are transported to these organelles by lipid-carrying proteins that extract a phospholipid molecule from one membrane and deliver it into another. Characteristics of mitochondrial genome The human mitochondrial DNA is circular, double-stranded structures that consist of base pairs carrying information of genes. They have a circular shape. A mitochondrion has multiple copies of its DNA, and a typical cell has many mitochondria; therefore, cells have many copies of mitochondrial DNA. Mitochondrial DNA have no histones. Mitochondrial DNA, unlike nuclear DNA, is inherited from the mother, while nuclear DNA is inherited from both parents. This is very helpful sometimes in determining how a person has a certain disorder in the family. New mitochondria arise from binary fission splitting. If all the mitochondria are removed from a cell, it cannot make new ones because there are no existing mitochondria to split. Vishal Kumar Evidences for the origin of mitochondria Structure and function of peroxisome (oxidative processes, catalase, peroxidase) Peroxisomes are enclosed in a single membrane and are. micrometer in diameter. They contain H​ O​ ​ ​ producing enzymes like oxidases and catalases as well as oxidative enzymes like peroxidase, Catalase, glycolic acid oxidase and some other enzymes. Proteins are selectively imported into peroxidases. Peroxisomes contain no DNA or ribosomes and have no means of producing proteins. Instead, all of these proteins are imported across the membranes. Peroxisomes are involved in the formation and decomposition of hydrogen peroxide and the word peroxisome is actually derived from hydrogen peroxide. Peroxisome contains oxidative enzymes, such as catalase, D-amino acid oxidase and uric acid oxidase. They use molecular oxygen to remove hydrogen atoms from a specific organic substrate R in an oxidative reaction. It produces hydrogen peroxide H​ O​ ​ ​ is a toxic byproduct of cellular metabolism. Catalases uses this H​ O​ ​ ​ in the peroxisome to oxidize other substrates like phenols, formic acid, formaldehyde, and alcohol. This reaction is important in liver and kidney cells where the peroxisomes detoxify various toxic substances that enter the blood. Peroxisomes are important for lipids metabolism. In humans, oxidation of fatty acids greater than carbons in length occurs in peroxisomes. A major function of peroxisomes is the breakdown of fatty acid molecules in a process called ​beta-oxidation​. In this process, the fatty acids are broken down into Acetyl-CoA. It is then transported to back to the cytosol for further use. Peroxisomes contain the first two enzymes required for the synthesis of plasmalogens myelin sheath. Peroxisomes also play important roles in cholesterol and bile acid synthesis, purine and polyamine catabolism, and prostaglandin metabolism. In plants, peroxisomes are required for photorespiration. Origin of peroxisome, protein transport (posttranslational transmembrane transport) into peroxisome Vishal Kumar Comparison of mitochondria and peroxisomes Mitochondria are double-membraned organelles, while peroxisomes are single-membraned. Peroxisomes do not contain DNA, while mitochondria do. Both the organelles grow and split. The main function of peroxisomes are the detoxifications of various toxic substances that enter the blood. The mitochondria are responsible for the energy metabolism, produce ATP and known as the powerhouse of the cell. 0. Cells in their social context. Extracellular matrix (ECM).​ ​ ​ECB​: , - OR ​ – , - 0 , 0 Fig. 0- ​.​ ​ lecture of DGCI th​ ​ week practice​ th​ Types of adhesion molecules (cadherins, selectins, Ig-like adhesion molecules) The cell junctions that hold an epithelium together by forming mechanical attachments are of three main types. Adherens junctions and desmosomes bind one epithelial cell to another, while hemidesmosomes bind epithelial cells to the basal lamina. All of these junctions provide mechanical strength by the same strategy: the proteins that form the cell adhesion span the plasma membrane and are linked inside the cell to cytoskeletal filaments. In this way, the cytoskeletal filaments are tied into a network that extends from cell to cell across the whole expanse of the epithelial sheet. Adherens junctions and desmosomes are both built around transmembrane proteins that belong to the cadherin family: a ​cadherin molecule in the plasma membrane of one cell binds directly to an identical cadherin molecule in the plasma membrane of its neighbor. Inside Vishal Kumar the cell, they are attached via linker proteins either actin filaments or keratin intermediate filaments. Such binding of like-to-like is called homophilic binding. In the case of cadherins, binding also requires that Ca be present in the extracellular medium hence the name. Immunoglobin-like adhesion molecules: usually homophilic binding Selectins: are heterophilic bindings. Ca dependent. Selectins have been implicated in several roles but they are especially important in the immune system by helping white blood cell homing and trafficking ECM-binding molecules (integrins, proteoglycans) If cells are to pull on the extracellular matrix and crawl over it, they must be able to attach to it. Cells do not attach well to bare collagen, therefore another protein fibronectin provides linkage. One part of the fibronectin binds to the collagen, while the other end attaches to a site on the cell on an integrin. Integrin is a receptor protein which spans the cell’s plasma membrane. When the extracellular domain of the integrin binds to fibronectin, the intracellular domain binds through a set of adaptor molecules to an actin filament inside the cell. Without this internal anchorage to the cytoskeleton, integrins would be ripped out of the flimsy lipid bilayer of the plasma membrane as the cell attempted to pull itself along the matrix Homophylic and heterophylic binding Homophilic binding: where adhesion molecules on one cell interact with identical molecules on another cell cadherins, immunoglobulin-like adhesion molecules Heterophilic binding: where an adhesion molecule on one cell functions as a receptor that binds to a different but specific molecule known as ligand on the other cell. selectins Structure and function of cell - cell junctions: tight junction (zonula occludens), belt desmosome (zonula adherens), spot desmosome (desmosome, macula adherens), and cell – extracellular junctions: focal contact (adhesion plaque), hemidesmosome Tight junctions (zonula occludens)​: The sealing function is served in vertebrates by tight junctions. These junctions seal neighboring cells together so that water-soluble molecules cannot easily leak between them. The tight junction is formed from proteins called claudins and occludins, which are arranged in strands along the lines of the junction to create the seal. Without tight junctions to prevent leakage, the pumping activities of absorptive cells such as those in the gut would be futile, and the composition of the extracellular fluid would become the same on both sides of the epithelium. Tight junctions also play a key part in maintaining the polarity of the individual epithelial Vishal Kumar cells in two ways. First, the tight junctions around the apical region of each cell prevents diffusion of proteins within the plasma membrane and so keeps the apical domain of the plasma membrane different from the basal or basolateral domain. Second, in many epithelia, the tight junctions are sites of assembly for the complexes of intracellular proteins that govern the apico-basal polarity of the cell interior. Adherens belt​: At an adherens junction, each cadherin molecule is tethered inside its cell, via several linker proteins, to actin filaments. Often, the adherens junctions form a continuous adhesion belt around each of the interacting epithelial cells; this belt is located near the apical end of the cell, just below the tight junctions. Bundles of actin filaments are thus connected from cell to cell across the epithelium. This network of actin filaments can contract, giving the epithelial sheet the capacity to develop tension and to change its shape in remarkable ways. By shrinking the apical surface of an epithelial sheet along one axis, the sheet can roll itself up into a tube. Alternatively, by shrinking its apical surface locally along all axes at once, the sheet can invaginate into a cup and eventually create a spherical vesicle by pinching off from the rest of the epithelium. Epithelial movements such as these are important in embryonic development. Desmosomes​: At a desmosome, a different set of cadherin molecules connects to keratin filaments the intermediate filaments found specifically in epithelial cells. Bundles of ropelike keratin filaments criss-cross the cytoplasm and are “spot-welded” via desmosome junctions to the bundles of keratin filaments in adjacent cells. This arrangement confers great tensile strength on the epithelial sheet and is characteristic of tough, exposed epithelia such as the epidermis of the skin. Hemidesmosomes: ​Blisters are a painful reminder that it is not enough for epidermal cells to be firmly attached to one another: they must also be anchored to the underlying connective tissue. As we noted earlier, the anchorage is mediated by integrins in the cells’ basal plasma membranes. The extracellular domains of these integrins bind to laminin in the basal lamina; inside the cell, the integrin tails are linked to keratin filaments, creating a structure that looks superficially like half a desmosome. These Vishal Kumar attachments of epithelial cells to the basal lamina beneath them are therefore called hemidesmosomes. Focal contact: ​Focal adhesions are integrin-containing, multi-protein structures that form mechanical links between intracellular actin bundles and the extracellular matrix or substrate in many cell types. Focal adhesions are in a state of constant flux: proteins associate and disassociate with it continually as signals are transmitted to other parts of the cell. They link cells to basal lamina. Structure and function of gap junction The final type of epithelial cell junction, found in virtually all epithelia and in many other types of animal tissues, serves a totally different purpose. In the electron microscope, this gap junction appears as a region where the membranes of two cells lie close together and exactly parallel, with a very narrow gap of nm between them. The gap, however, is not entirely empty; it is spanned by the protruding ends of many identical, transmembrane protein complexes that lie in the plasma membranes of the two apposed cells. These complexes, called connexons, are aligned end-to-end to form narrow, water-filled channels across the two plasma membranes The channels allow inorganic ions and small, water-soluble molecules up to a molecular mass of about daltons to move directly from the cytosol of one cell to the cytosol of the other. This creates an electrical and a metabolic coupling between the cells. Gap junctions between cardiac muscle cells, for example, provide​ ​the electrical coupling that allows electrical waves of excitation to spread synchronously through the heart, triggering the coordinated contraction of the cells that produces each heartbeat.. Cytoskeleton.​ ​ ECB: OR - ​ ​ lecture of DGCI​ - th​ Vishal Kumar Microtubules (tubulin, polymerization, depolymerization, polarity of microtubules, dynamism, MAPs, motor proteins: dyneins, kinesins) General: ​Microtubules have a crucial organizing role in all eukaryotic cells. These long and relatively stiff hollow tubes of protein can rapidly disassemble in one location and reassemble in another. In a typical animal cell, microtubules grow out from a small structure near the center of the cell called the centrosome. Extending out toward the cell periphery, they create a system of tracks within the cell, along which vesicles, organelles, and other cell components can be transported. These cytoplasmic microtubules are the part of the cytoskeleton mainly responsible for transporting and positioning membrane-enclosed organelles within the cell and for guiding the intracellular transport of various cytosolic macromolecules. When a cell enters mitosis, the cytoplasmic microtubules disassemble and then reassemble into an intricate structure called the mitotic spindle. The mitotic spindle provides the machinery that will segregate the chromosomes equally into the two daughter cells just before a cell divides. Microtubules can also form stable structures, such as rhythmically beating cilia and flagella. These hairlike structures extend from the surface of many eukaryotic cells, which use them either to swim or to sweep fluid over their surface. The core of a eukaryotic cilium or flagellum consists of a highly organized and stable bundle of microtubules. Tubulin: ​Microtubules are built from subunits molecules of tubulin each of which is itself a dimer composed of two very similar globular proteins called α-tubulin and β-tubulin, bound tightly together by noncovalent interactions. The tubulin dimers stack together, again by noncovalent bonding, to form the wall of the hollow cylindrical microtubule. This tubelike structure is made of parallel protofilaments, each a linear chain of tubulin dimers with α- and β-tubulin alternating along its length. One end of the microtubule, thought to be the β-tubulin end, is called its plus end, and the other, the α-tubulin end, its minus end. This gives the polarity of microtubule as a whole. In a concentrated solution of pure tubulin in a test tube, tubulin dimers will add to either end of a growing microtubule. However, they add more rapidly to the plus end than to the minus end, which is why the ends were originally named this way not because they are electrically charged. The polarity of the microtubule the fact that its structure has a definite direction, with the two ends being chemically and functionally distinct is crucial, both for the assembly of microtubules and for their role once they are formed. If microtubules had no polarity, they could not guide intracellular transport, for example. MTOC, -tubulin Vishal Kumar Inside cells, microtubules grow from specialized organizing centers that control the location, number, and orientation of the microtubules. In animal cells, for example, the centrosome​ which is typically close to the cell nucleus when the cell is not in mitosis organizes an array of microtubules that radiates outward through the cytoplasm. The centrosome consists of a pair of centrioles, surrounded by a matrix of proteins. The centrosome matrix includes hundreds of ringshaped structures formed from a special type of tubulin, called γ-tubulin, and each γ-tubulin ring complex serves as the starting point, or nucleation site, for the growth of one microtubule. The αβ-tubulin dimers add to each γ-tubulin ring complex in a specific orientation, with the result that the minus end of each microtubule is embedded in the centrosome, and growth occurs only at the plus end that extends into the cytoplasm. Dynamic instability: ​The dynamic instability of microtubules stems from the intrinsic capacity of tubulin dimers to hydrolyze GTP. Each free tubulin dimer contains one GTP molecule tightly bound to β-tubulin, which hydrolyzes the GTP to GDP shortly after the dimer is added to a growing microtubule. This GDP remains tightly bound to the β-tubulin. When polymerization​ ​is proceeding rapidly, tubulin dimers add to the end of the microtubule faster than the GTP they carry is hydrolyzed. As a result, the end of a rapidly growing microtubule is composed entirely of GTP-tubulin dimers, which form a “GTP cap.” GTP-associated dimers bind more strongly to their neighbors in the microtubule than do dimers that bear GDP, and they pack together more efficiently. Thus the microtubule will continue to grow Figure A. Because of the randomness of chemical processes, however, it will occasionally happen that the tubulin dimers at the free end of the microtubule will hydrolyze their GTP before the next dimers are added, so that the free ends of protofilaments are now composed of GDP-tubulin. These GDP-bearing dimers associate less tightly, tipping the balance in favor of disassembly Figure B. Because the rest of the microtubule is composed of GDP-tubulin, once depolymerization has started, it will tend to continue; the microtubule starts to shrink rapidly and continuously and may even disappear. Motor proteins: ​This saltatory movement moving for a short period, stopping, and then moving again is much more sustained and directional than the continual, small, Brownian movements caused by random thermal motions. Saltatory movements can occur along either microtubules or actin filaments. In both cases, the movements are driven by motor proteins, which use the energy derived from repeated cycles of ATP hydrolysis to travel steadily along the microtubule or actin filament in a single direction. Because the motor proteins also attach to other cell components, they can transport this cargo along the filaments. Vishal Kumar The motor proteins that move along cytoplasmic microtubules, such as those in the axon of a nerve cell, belong to two families: the ​kinesins​ generally move toward the ​plus​ end of a microtubule, the dyneins move toward the minus end. Both kinesins and dyneins are generally dimers that have two globular ATP-binding heads and a single tail. The heads interact with microtubules in a stereospecific manner, so that the motor protein will attach to a microtubule in only one direction. The tail of a motor protein generally binds stably to some cell component, such as a vesicle or an organelle, and thereby determines the type of cargo that the motor protein can transport. The globular heads of kinesin and dynein are enzymes with ATP-hydrolyzing ATPase activity. This reaction provides the energy for driving a directed series of conformational changes in the head that enable it to move along the microtubule by a cycle of binding, release, and rebinding to the microtubule. Cilia and flagellum: ​Cilia are hairlike structures about. μm in diameter, covered by plasma membrane, that extend from the surface of many kinds of eukaryotic cells; each cilium contains a core of stable microtubules, arranged in a bundle, that grow from a cytoplasmic basal body, which serves as an organizing center. Cilia beat in a whiplike fashion, either to move fluid over the surface of a cell or to propel single cells through a fluid. Cilia has a x structure. We can find cilia in trachea and fallopian tube cells. The flagella singular flagellum that propel sperm and many protozoa are much like cilia in their internal structure but are usually very much longer. They are designed to move the entire cell, rather than moving fluid across the cell surface. Flagella propagate regular waves along their length, propelling the attached cell along. Sperm cells have flagella. A cross section through a cilium shows nine doublet microtubules arranged in a ring around a pair of single microtubules. This “ ” array is characteristic of almost all eukaryotic cilia and flagella Actin or (micro)filaments (polymerization, depolymerization, polarity) General: ​Actin filaments, polymers of the protein actin, are present in all eukaryotic cells and are essential for many of the cell’s movements, especially those involving the cell surface. Without actin filaments, for example, an animal cell could not crawl along a surface, engulf a large particle by phagocytosis, or divide in two. Like microtubules, many actin filaments are unstable, but by associating with other proteins they can also form stable structures in cells, such as the contractile apparatus of muscle cells. Actin filaments interact with a large Vishal Kumar number of actin-binding proteins that enable the filaments to serve a variety of functions in cells. Depending on which of these proteins they associate with, actin filaments can form stiff and stable structures, such as the microvilli on the epithelial cells lining the instestine or the small contractile bundles that contract. Actin filaments are thinner, more flexible, and usually shorter than microtubules. There are, however, many more of them, so that the total length of all the actin filaments in a cell is generally many times greater than the total length of all of the microtubules. Unlike intermediate filaments and microtubules, actin filaments rarely occur in isolation in the cell; they are generally found in cross-linked bundles and networks, which are much stronger than the individual filaments. (De) polymerization: Although actin filaments can grow by the addition of actin monomers at either end, like microtubules, their rate of growth is faster at the plus end than at the minus end. A naked actin filament, like a microtubule without associated proteins, is inherently unstable, and it can disassemble from both ends. In living cells, free actin monomers carry a tightly bound nucleoside triphosphate, in this case ATP. The actin monomer hydrolyzes its bound ATP to ADP soon after it is incorporated into the filament. As with the hydrolysis of GTP to GDP in a microtubule, hydrolysis of ATP to ADP in an actin filament reduces the strength of binding between the monomers, thereby decreasing the stability of the polymer. Thus in both cases, nucleotide hydrolysis promotes depolymerization, helping the cell to disassemble its microtubules and actin filaments after they have formed. If the concentration of free actin monomers is very high, an actin filament will grow rapidly, adding monomers at both ends. At intermediate concentrations of free actin, however, something interesting takes place. Actin monomers add to the plus end at a rate faster than the bound ATP can be hydrolyzed, so the plus end grows. At the minus end, by contrast, ATP is hydrolyzed faster than new monomers can be added; because ADP-actin destabilizes the structure, the filament loses subunits from its minus end at the same time as it adds them to the plus end Figure. In as much as an individual monomer moves through the filament from the plus to the minus end, this behavior is called treadmilling. Treadmilling involves a simultaneous gain of monomers at the plus end of an actin filament and loss at the minus end: when the rates of addition and loss are equal, the filament remains the same size. Vishal Kumar ABP: structural and motor proteins There are a great many actin-binding proteins in cells. Most of these bind to assembled actin filaments rather than to actin monomers and control the behavior of the intact filaments. Actin-bundling proteins, for example, hold actin filaments together in parallel bundles in microvilli; others cross-link actin filaments together in a gel-like meshwork within the cell cortex the specialized layer of actin-filament-rich c