LAB EVALUATION OF CELL DISORDERS

2) Transport across cell membranes

Ola H. Elgaddar

MBChB, MSc, MD, CPHQ, LGBSS Lecturer of Chemical Pathology

Medical Research Institute Alexandria University

Ola.elgaddar@alexu.edu.eg

Sources:

1. Grey's anatomy, 39th ed, 2008

2. Ganong physiology, 23rd ed,2010

3. Robbins pathology, 7th ed, 2005

4. Kumar clinical medicine,6th ed,2005

5. Molecular cell biology, 5th ed, 2003

- The cell performs many functions like support, transport, communication, recognition and acts as selective barriers. All these functions are achieved through movement across cell membranes.

- There are several mechanisms of transport across cellular membranes. Pathways include passive and active transport.

- Small, non-polar molecules (including O2 and N2) and small uncharged polar molecules such as CO2 diffuse across the lipid membranes of cells (Passive simple diffusion)

-However, the membranes have very limited permeability to other substances. Instead, they cross the membranes by endocytosis and exocytosis or by passage through highly specific transport proteins. These are transmembrane proteins that are either ion channels , ATP-powered pumps or carrier proteins.

- The limited permeability applies even to water, with simple diffusion being supplemented throughout the body with various water channels (aquaporins).

A)Ion channels:

-Channel proteins transport water or specific types of ions and hydrophilic small molecules down their concentration or electric potential gradients.

- Such protein-assisted transport sometimes is referred to as facilitated diffusion.

- Channel proteins form a hydrophilic passageway across the membrane through which multiple water molecules or ions move simultaneously, single file at a very rapid rate.

- Some ion channels are open much of the time; these are referred to as non-gated channels.

- Most ion channels, however, open only in response to specific chemical or electrical signals; these are referred to as gated channels.

1-Cell structure.pptx

-Though ion channels shuttle between a closed state and an open state, many ions can pass through an open channel without any further conformational change. For this reason, channels are characterized by very fast rates of transport, up to 108 ions per second.

- There are ion channels specific for K+, Na+, Ca2+, and Cl–, as well as channels that are nonselective for cations or anions.

Ways in which ion channels form pores:

(A) K+ channels are tetramers, with each protein subunit forming part of the channel.

(B) In ligand-gated cation and anion channels such as the acetylcholine receptor, five identical or very similar subunits form the channel.

(C) Cl– channels are dimers, with an intracellular pore in each subunit.

(D) Aquaporin water channels are tetramers with an intracellular channel in each subunit.

B) Pumps (ATP-powered pumps):

- They are ATPases that use the energy of ATP hydrolysis to move ions or small molecules across a membrane against a chemical concentration gradient or electric potential or both.

- This process, referred to as active transport, is an example of a coupled chemical reaction. In this case, transport of ions or small molecules “uphill” against an electrochemical gradient, which requires energy, is coupled to the hydrolysis of ATP, which releases energy.

Examples of ATPases:

Na, K ATPase: known as the Na, K pump.

H, K ATPases: in the gastric mucosa and the renal tubules.

Ca2+ATPase: pumps Ca2+ out of cells.

Na, K -ATPase:

- It catalyzes the hydrolysis of ATP to ADP and uses the energy to extrude three Na+ from the cell and take two K+ into the cell for each molecule of ATP hydrolyzed (Active transport)

- It is an electrogenic pump in that it moves three positive charges out of the cell for each two that it moves in, and it is therefore said to have a coupling ratio of 3:2

- It is found in all parts of the body. Its activity is inhibited by ouabain and related digitalis glycosides used in the treatment of heart failure.

../DATA/Sodium Potassium Pump.flv

C) Carrier proteins

(Transporters):

- They bind ions and other molecules and then change their configuration, moving the bound molecule from one side of the cell membrane to the other.

Two types of transporters:

1) Uniporters

2) Co-transporters

1) Uniporters:

transport a

single type of

molecule

down its

concentration

gradient via

facilitated diffusion.

Ex: GLUT 1

2) Co-transporters:

-Couple the movement of one type of ion or molecule against its concentration gradient (the driven substrate) with the movement of one or more different ions down its concentration gradient (the driving substarte).

(Secondary active transport)

- The direction of transmembrane movement of the driven substrate can be either the same as (symport), or opposite to (antiport), that of the driving substrate.

The luminal membranes of mucosal cells in the small intestine contain a symport that transports glucose into the cell only if Na+ binds to the protein and is transported into the cell at the same time.

In the heart, Na, K ATPase indirectly affects Ca2+ transport. An antiport in the membranes of cardiac muscle cells normally exchanges intracellular Ca2+ for extracellular Na+

- Like ATP pumps, co-transporters mediate coupled reactions in which an energetically unfavorable reaction (i.e., uphill movement of molecules) is coupled to an energetically favorable reaction.

- Note, however, that the nature of the energy-supplying reaction driving active transport by these two classes of proteins differs. ATP pumps use energy from hydrolysis of ATP, whereas co-transporters use the energy stored in an electrochemical gradient.

- ATP-powered pumps and transporters undergo a cycle of conformational changes exposing a binding site (or sites) to one side of the membrane in one conformation and to the other side in a second conformation. Because each cycle results in movement of only one (or a few) substrate molecules, these proteins are characterized by relatively slow rates of transport ranging from 10 to 104 ions or molecules per second.

Transport across cell membranes

- Secreted proteins, lipid, small molecules and other cellular products destined for export from the cell are transported to the plasma membrane in small vesicles released from the trans face of Golgi.

- This pathway is either constitutive, in which transport and secretion occur more or less continuously, or it is regulated by external signals, as in the control of salivary secretion by autonomic neural stimulation.

-Exocytosis is achieved by fusion of the secretory vesicular membrane with the plasma membrane and release of the vesicle contents into the extracellular domain.

- Excess plasma membrane generated by vesicle fusion during exocytosis is rapidly removed by concurrent endocytosis.

- The process of endocytosis involves the internalization of vesicles derived from the plasma membrane.

- The vesicles may contain: engulfed fluids and solutes from the extracellular interstitial fluid (pinocytosis);larger macromolecules, often bound to surface receptors (receptor-mediated endocytosis); particulate matter, including microorganisms or cellular debris (phagocytosis).

1 - Pinocytosis generally involves small fluid-filled vesicles and is a marked property of capillary endothelium, e.g. where vesicles containing nutrients and oxygen dissolved in blood plasma are transported from the vascular lumen to the endothelial basal plasma membrane.

Interstitial fluid containing dissolved carbon dioxide is also taken up by pinocytosis for simultaneous transportation across the endothelial cell wall in the opposite direction, for release into the bloodstream by exocytosis. This shuttling of pinocytotic vesicles is

also termed transcytosis.

N.B: Transcytosis is a process used by some cells for the apical-basolateral sorting of certain membrane proteins. This process of transcellular transport, which combines endocytosis and exocytosis, also can be employed to import an extracellular ligand from one side of a cell, transport it across the cytoplasm, and secrete it from the plasma membrane at the opposite side.

2 - Receptor-mediated endocytosis, also known as clathrin-dependent endocytosis, is initiated at specialized regions of the plasma membrane known as clathrin-coated pits.

Clathrin is a protein which plays a major role in the formation of coated vesicles. It forms a triskelion shape (a figure with three bent legs), which is composed of three clathrin heavy chains and three light chains. The legs are joined at a central trimerization domain (txd). When the triskelion interact they form a polyhedral lattice which surrounds the vesicle.

- Adaptors represent a diverse group of proteins recognizing different classes of cargo receptor.

-The best characterized are a family of closely related proteins called the adaptor proteins (APs) comprising AP1, AP2, AP3 and AP4.

- Each of these four classes are localized to different intracellular compartments and vary in their receptor specificity

-Clathrin assembles onto the adaptors to form the outer layer of the coat.

-It acts to stabilise the curvature introduced into the growing pit whilst increasing its deformation until the entire region invaginates to form a closed vesicle.

3 - Phagocytosis:

- Phagocytosis is a property of many cell types, but is most efficient in cells specialized for this activity.

-The professional phagocytes of the body belong to the monocyte lineage of haemopoietic cells, in particular the tissue macrophages.

-Other effective phagocytes are neutrophil granulocytes and most dendritic cells, which are also of haemopoietic origin.

-Phagocytosis is a triggered process, initiated when a phagocytic cell binds to a particle or organism, often through a process of molecular recognition.

-Typically, a pathogenic microorganism may first be coated by antibodies, which are bound in turn by receptors for the Fc portion of the antibody molecule expressed by macrophages and neutrophils.

-This triggers the production of large pseudopodia, which engulf the organism and internalize it.

-The process appears to depend on actin-myosin-based cellular motility and, unlike receptor-mediated endocytosis, it is energy dependent.

- Inside the cell, the phagosome fuses with lysosomes, which degrade its contents.

-Lysosomes are membrane-bound intracellular organelles that contain a variety of hydrolytic enzymes, including acid phosphatase, glucuronidase, sulfatase, ribonu-clease, and collagenase.

- These enzymes are synthesized in the rough endoplasmic reticulum and then packaged into vesicles in the Golgi apparatus.

- Lysosomes (primary) fuse with membrane-bound vacuoles that contain material to be digested, forming secondary lysosomes or phagolysosomes

Lysosomes are involved in the breakdown of phagocytosed or endocytosed materials in one of two ways: heterophagy or autophagy.

Heterophagy is the process of lysosomal digestion of materials ingested from the extracellular environment.

- Extracellular materials are taken up by cells through the general process of endocytosis (Pinocytosis or phagocytosis).

-Heterophagy is most common in the "professional" phagocytes, such as neutrophils and macrophages, although it may also occur in other cell types.

- Examples of heterophagocytosis include the uptake and digestion of bacteria by neutrophils

Autophagy refers to lysosomal digestion of the cell's own components.

- In this process, intracellular organelles and portions of cytosol are first sequestered from the cytoplasm in an autophagic vacuole formed from ribosome-free regions of the rough endoplasmic reticulum.

- The vacuole fuses with lysosomes or Golgi elements to form an autophagolysosome.

Autophagy is a common phenomenon involved in the removal of damaged organelles during cell injury and the cellular remodeling of differentiation, and it is particularly pronounced in cells undergoing atrophy induced by nutrient deprivation or hormonal involution.

-The enzymes in lysosomes are capable of degrading most proteins and carbohydrates, but some lipids remain undigested.

- Lysosomes with undigested debris may persist within cell as residual bodies or may be extruded.

Lipofuscin pigment granules represent undigested material derived from intracellular lipid peroxidation.

- Certain indigestible pigments, such as carbon particles inhaled from the atmosphere or inoculated pigment in tattoos, can persist in phagolysosomes of macrophages for decades.

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