Lesson 2.2 -Membrane Transport 2023-2024 PDF

CELL BIOLOGY AND HUMAN GENETICS Ve más allá Academic Year 2023-2024 DAVID BALLESTEROS PLAZA Department of Pre-clinical Dentistry (Building A) E-mail address: davidalberto.ballesteros @universidadeuropea.es Ve más allá Academic Year 2023-2024 LESSON 2 PART II MEMBRANE TRANSPORT MEMBRANE PERMEABILITY...

CELL BIOLOGY AND HUMAN GENETICS Ve más allá Academic Year 2023-2024 DAVID BALLESTEROS PLAZA Department of Pre-clinical Dentistry (Building A) E-mail address: davidalberto.ballesteros @universidadeuropea.es Ve más allá Academic Year 2023-2024 LESSON 2 PART II MEMBRANE TRANSPORT MEMBRANE PERMEABILITY BIOLOGICAL MEMBRANES ACT AS SELECTIVE BARRIERS Biological membranes regulate the transport of different molecules : Between extracellular and intracellular spaces Between different intracellular compartments. All cellular membranes, both plasma and organelle, must serve not only as barriers, but also as exchange platform selectively transporting molecules and ions from one side of the membrane to the other. PRINCIPLES OF MEMBRANE TRANSPORT Ion concentrations are very different Inside and Outside a Cell These differences are crucial for all cell´s survival and function. Inorganic ions such as Na+, K+, Ca2+, Cl- and H+ are essential for ATP production and for nerve cell activity. PHOSPHOLIPID BILAYER HAVE PERMEABILITY DEPENDING ON MOLECULE SIZE AND POLARITY (OR CHARGE) Pure Lipid Bilayers Are impermeable to large watersoluble molecules and ions. Water and small nonpolar molecules can move accross the bilayer by simple diffusion. TRANSPORT MECHANISMS FOR DIFFERENT MOLECULES (low and high weight) TRANSPORT OF LOW MOLECULAR WEIGHT MOLECULES PASSIVE TRANSPORT SIMPLE DIFUSSION FACILITATED DIFUSSION ACTIVE TRANSPORT TRANSPORT OF HIGH MOLECULAR WEIGHT MOLECULES ENDOCYTOSIS EXOCYTOSIS TRANSCYTOSIS Na+-K+ Pump Na+ Glucosesymport PINOCYTOSIS PHAGOCYTOSIS RECEPTOR- MEDIATED ENDOCYTOSIS POLAR OR CHARGED MOLECULES CROSS BIOLOGICAL MEMBRANES BY PASSIVE OR ACTIVE TRANSPORT Movement DOWN the concentration gradient DOES NOT REQUIRE ENERGY Movement AGAINST the concentration gradient (UP) REQUIRES ENERGY and specialized transport proteins GRADIENTS AND MEMBRANE TRANSPORT The movement of charged solutes (ions) accross a membrane involves two gradients :  The chemical gradient (concentration gradient)  The membrane potential (charge gradient) Both gradients contribute to generate the driving force that moves ions inside ELECTROCHEMICAL GRADIENT In this case, the concentration gradient favors the movement up but the Electric Potential opposes it. Ions will diffuse out but the driving force will be smaller MEMBRANE TRANSPORT PROTEINS (MTP) Membrane transport proteins (MTP) Fall into Three Categories : ATP-Powered PUMPS, TRANSPORTERS and CHANNELS All transport proteins are transmembrane proteins containing multiple membranespanning segments that are generally α helices. These proteins form a protein-lined pathway across the membrane that allows movement of hydrophylic substances without contacting the hydrophobic interior of the membrane. PASSIVE TRANSPORT : SIMPLE DIFFUSION Transport of small uncharged molecules down their concentration gradient  It does not require transport proteins  It does not require Energy  It is a non-selective process Urea PASSIVE TRANSPORT : FACILITATED DIFFUSION Transport of larger polar molecules or ions down their concentration gradient Channels Transporters or Carriers  It does not require Energy  It requires transmembrane proteins : Permanently opened  Channels Regulated  Porins  Transporter or Carrier Proteins Porins IONIC CHANNELS OR PORES -Channels are selective -Channels can be regulated -By voltage, -by ligand Potassium channel - mechanically -Do not saturate PORINS Bigger than channels They allow the passage of polar substances Substance selective Permanently open AQUAPORINS, MALTOPORINE Do not saturate CARRIERS Substance selective The binding of the substance to be transported induces a change in conformation and opening to the other side (GLUT) They saturate 13 CHANNELS WITH REGULATED OPENING Channel proteins form pores in the membrane through which the transported molecule circulates without coming in contact with the hydrophobic region of the bilayer. Ion Channels are usually very selective Channels can be permanently opened or regulated Ion Channels with regulated opening : they are usually closed and open in response to a chemical , mechanical or electrical signal. (A)Electrical signal: Activated by voltage (depolarization) (B) and (C) Chemical signal : Activated by ligand (D) Mechanical signal: Mechanically activated (Release of calcium in sarcoplasmic reticulum) Activated by ligand NEUROTRANSMITTERS Acetilcholin Receptor Acerilcholin  GABA Receptors (canales de Cl-) The binding of Ach induces a conformational change Nicotinic receptor Na+ Closed channel The binding of acetylcholine to a site of the extracellular domain of R induces conformational changes that open the gate of the channel, allowing the passage of + charged ions. The channel have negatively charged amino acids that prevent negative ion flow. Na+ Open cjannel 15 Voltage gated channels + + + + - - - - + + + - - - - + + + + + + + + + + The regulation of the opening by voltage depends on a transmembrane alpha helix, which contains multiple charged aas +. The depolarization of the mb induces the movement of these positive charges towards the outside of the mb, which alters the position of this transmembrane segment and induces the opening of the channel These types of channels are very selective + + 16 Mechanically regulated channels In the myocyte, the action potential travels through the T tubules and activates DHP (Dihydropyridine) receptors, which undergo a conformational change. The R DHP and the Ca2 + channels face each other, so that the conformational change in the R DHP causes the opening of the Ca2 + channel and the exit of Ca2 + present in the mb of the sarcoplasmic reticulum. Ca2 + exits massively into the cytoplasm of the muscle fiber and regulates muscle contraction DHP: dihidropiridina PORINS  They allow the free passage of specific polar molecules.  They cannot be saturated.  Examples : aquaporins , maltoporin (in bacteria) AQUAPORINS are water channel proteins that facilitate the transport of water through different cell membranes. Water molecules go through the channel in a single line These transmembrane protein channels form a pore of a size that only allows the passage of water molecules. They are present in animal and plant cells Transport of water molecules through aquaporins is faster than simple diffusion. TRANSPORTERS OR CARRIERS  Carrier proteins or transporters bind SPECIFIC molecules on one side of the membrane, undergo a CONFORMATIONAL CHANGE and release the molecule on the other side.  They move molecules through the membrane at a much lower rate than channels.  Glucose transport to the cell´s interior is a well studied example of transporter-mediated facilitated diffusion, since most cells in our body are exposed to higher glucose concentrations in the extracellular medium.  The conformational change in the glucose transporter is induced by the binding of glucose and is reverted once glucose is released on the other side of the membrane. Humans have at least five related Glucose transporter. These isoforms ( GLUT 1 to GLUT 5 ) are expressed in different tissues and show different kinetics and regulation. ACTIVE TRANSPORT  Transport of ions or other molecules AGAINST THEIR CONCENTRATION GRADIENT  It is energetically unfavourable and therefore REQUIRES ENERGY INPUT. Depending on the source of energy, we can distinguish :  Primary Active Transport : uses energy fom ATP hydrolysis  Secondary Active Transport : uses energy from an ion concentration gradient or an electrical gradient. Primary Active Transport Secondary Active Transport Concentration Gradient COUPLED TRANSPORT Three Different Movements PRIMARY ACTIVE TRANSPORT: THENa+-K+ PUMP This protein transporter uses the energy derived from ATP hydrolysis to pump 3 molecules of Na+ from the cytosol to the extracellular medium and 2 molecules of K+ from the exterior to the cytosol. In both cases the flow is against gradient. The Na+-K+ pump is a transporter and an enzyme, typically accounting for 30% of total cellular ATP consumption. Na+ ions bind to high affinity sites of the pump inside the cell (1). This binding stimulates ATP hydrolysis and phosphorylation of the pump (2), which in turn induces a conformational change leading to Na+ release on the outside of the cell (3). At the same time, high affinity sites for K+ are exposed on the cell surface (4). The binging of extracellular K+ ions stimulates dephosphorylation of the pump (4) which induces a second conformational (5) change leading to the release of K+ on the cytosolic side (6). Na+ Na+ Na+ Na+ Na+ Na+ Na+ Na+ P ATP P ADP K+ K+ K+ K+ P P K+ K+ Na+ PRIMARY ACTIVE TRANSPORT The active transport of Na+, K+ and other ions across the plasma membrane is essential to maintain the intracellular ionic composition of cells and to keep ionic concentrations different in the intra- and extracellular medium. This, in turn, helps maintain the Osmotic Balance in animal cells Ion Intracellular (mM) Extracellular (mM) Na+ 5-15 145 K+ 140 5 Mg++ 30 1-2 Ca++ 1-2 2.5-5 Cl- 4 110 ACTIVE SECONDARY TRANSPORT In this kind of active transport the energy does not come directly from ATP but from an electrochemical gradient generated previously by primary active transport (for example, the Na+ gradient generated by the Na+-K+ pump). The Na+-Glucose Symporter is a good example of active secondary transport. Na+-Glucose Symporter Extracellular Space Na+ Concentration Gradient Cytosol 2 GLUCOSE UPTAKE IN EPITHELIAL INTESTINAL CELLS Because the Glucose concentration inside these epitelial cells is higher than the concentratrion in the intestinal lumen, Glucose absortion requires an active transport. In this case, the transport of Glucose against its concentration gradient is coupled to the transport of 2 Na+ ions down their electrochemical gradient. All 3 elements are transported simultaneously : SYMPORT. HIGH [Glucose] lumen H C Locate in the figure B D High glucose concentration Low glucose concentration Sodium / potassium pump Sodium / glucose symporter GLUT transporter Active sodium transport Secondary active glucose transport Passive glucose transport F E G capilar A 27 Simple diffusion (nonpolar compounds; down concentration gradient. Facilitated diffusion through carrier (in favor of electrochemical gradient) Primary active transport (up electrochemical gradient) Facilitated diffusion through ion channel (down the electrochemical gradient) Secondary active transport (againts electrochemical gradient driven by the ion moving down its gradient) SUMMARY OF THE FOUR MECHANISMS BY WHICH SMALL MOLECULES AND IONS ARE TRANSPORTED ACROSS CELLULAR MEMBRANES. TRANSPORT OF LARGE MOLECULES ACCROSS CELLULAR MEMBRANES TRANSPORT OF HIGH MOLECULAR WEIGHT MOLECULES ENDOCYTOSIS EXOCYTOSIS TRANSCYTOSIS PINOCYTOSIS PHAGOCYTOSIS RECEPTOR- MEDIATED ENDOCYTOSIS  Cells secrete macromolecules by EXOCYTOSIS  Cells take in macromolecules by ENDOCYTOSIS EXOCYTOSIS ENDOCYTOSIS In both processes , macromolecules are contained within VESICLES so that they do not mix with other components of the cytosol. Both EXOCYTOSIS and ENDOCYTOSIS depend on MEMBRANE FUSION. Membrane fusion occurs when two separate lipid membranes merge into a single continuous bilayer. ENDOCYTOSIS, EXOCYTOSIS, PINOOCYTOSIS, AND PHAGOCYTOSIS REQUIRE CLATRIN-COATED VESICLES Clathrin: Protein whose main function is to coat intracellular vesicles trisquelión Clathrin monomer is made up of 3 heavy chains and 3 light chains, which make up a structure called 'triskelion' Triskelions self-assemble into flat structures with a hexagonal arrangement, with which they cover the vesicles 32 TRANSPORT OF HUGE MOLECULES : ENDOCYTOSIS Process by which cells take in materials through an invagination of the plasma membrane that surrounds the ingested material in a membrane enclosed vesicle. CLASSIFICATION: Pinocytosis Phagocytosis Receptor-mediated endocytosis The “ingested” particles vary from macromolecules to complete cells. PINOCYTOSIS (Cell Drinking) Uptake of extracellular fluid and its content. It is a property of all eukaryotic cells. It involves the formation of invaginations in the cell membrane, that close and break off to form fluid-filled vacuoles in the cytoplasm. Most cells carry out pinocytosis. It is a non-selective, indiscriminate process. PHAGOCYTOSIS (Cell Eating) It involves the ingestion of large particles, such as microorganisms and cell debris, or even intact cells, via large vesicles called phagosomes (generally >250 nm in diameter) that will later fuse with lysosomes, forming phagolysosomes. ROLE OF PHAGOCYTOSIS : Nutrition : in unicelular organisms such as amoebas. Protection: in multicellular organisms. Defense against invading microorganims. Scavenging: multicellular organisms use phagocytosis to eliminate aged or damaged cells. In mammals phagocytosis is mostly carried out by macrophages and neutrophils. Macrophages in our spleen and liver dispose of more tan 1011 aged blood cells every day !!!! PHAGOCYTOSIS Phagocytosis by a White Blood Cell Scanning electron micrograph showing a macrophage engulfing a pair of red blood cells. RECEPTOR MEDIATED ENDOCYTOSIS Selective uptake of specific macromolecules (cholesterol, vitamin B12, iron virus) to be internalized. The macromolecules first need to bind to specific receptors. Very efficient and specific system. Ligand Receptor-Ligand Complex Endocytic Vesicle Clathrin These endocytic vesicles are coated with CLATHRIN, a protein made up of 3 heavy chains and 3 light chains assembled into a Triskelion structure. Plasma membrane TRANSPORT OF CHOLESTEROL THROUGH RECEPTOR MEDIATED ENDOCYTOSIS Cholesterol is extremely insoluble in water and cannot “travel” free in the bloodstream Cholesterol is transported through the bloodstream in lipoprotein particles called lowdensity lipoproteins, or LDL LDL particle outer surface: Amphipathic lipids and proteins 500 molecules of cholesterol 800 molecules of phospholipid 1 apoprotein B100 LDL particles bind to specific receptors located on the cell surface and the receptor–LDL complexes are ingested by receptor-mediated endocytosis and delivered to endosomes. The receptor is then recycled to the plasma membrane while LDL particles are transported to lysosomes where cholesterol is released for use by the cell. LDL Hydrophobic interior : 1500 molecules of cholesteryl esters CHOLESTEROL RECEPTOR MEDIATED ENDOCYTOSIS acid EXPORT OF LARGE MOLECULES : EXOCYTOSIS Process through which most molecules are secreted from a eukaryotic cell. These molecules (hormones, enzymes, mucus, etc..) and waste products are packaged in membrane-enclosed vesicles that fuse with the plasma membrane, releasing their contents to the extracellular medium. It is also be used to deliver proteins or lipids to the plasma membrane. Cytoskeleton Components such as actin are important in this process EXOCYTOSIS It is needed for :  SECRETION of proteins like enzymes, peptide hormones and antibodies.  TURNOVER of plasma membrane  ANTIGEN PRESENTATION during the immune response  RECYCLING of internalized plasma membrane receptors  DELIVERY of integral membrane proteins to the plasma membrane  RELEASE OF NEUROTRANSMITTER from presynaptic neurons EXOCYTOSIS COULD BE CONSTITUTIVE OR REGULATED RELEASE OF NEUROTRANSMITTERS. \. IN PRESYNAPTIC VESICLES TRANSCYTOSIS  Transcytosis is the process by which large molecules can be transferred across the cell to the opposite domain of the plasma membrane. For example in an epithelial cell from the basal membrane to the apical membrane.  It involves endocytosis followed by exocytosis.  The vesicle content is not modified during transport.  It is very common in endothelial cells for transfering large proteins from the blood to extracellular fluids. NON CLATRIN-COATED VESICLES : CAVEOLAE The endocytic vesicles in transcytosis are not clathrin-coated vesicles : they derive from CAVEOLAE. Caveolae are small invaginations of the plasma membrane (50-100 nm) that function in the uptake of selected molecules. Endocytic caveolae do not fuse with lysosomes. Caveolae are coated with CAVEOLIN, an integral membrane protein that forms dimers and is frequently found on the cytosolic side of lipid rafts. When several caveolin dimers are concentrated in a small region, they force a curvature in the lipid bilayer, forming a caveola.