Chapter 19- Cell junctions and the ECM

Question 1

Epithelial tissue

Answer 1

This type of tissue makes up the linings of different parts of the body, like the linings of the gut, airway, and skin. These linings are composed of many different cells that are joined by cell-cell junctions. There is some ECM called the basal lamina (or basement membrane) that underlies the epithelial tissue

Question 2

Connective tissue

Answer 2

Bone and tendons are examples, but connective tissue makes up the protective covering of other tissues and is found throughout the body. A lot of ECM is produced in connective tissue, and the cells are distributed sparsely. There are some cell-cell attachments, but many cell-ECM attachments

Question 3

Epithelial tissue structure

Answer 3

In the GI tract, epithelial tissue is made of columnar cells with microvilli on the apical surface. All of these cells are held together by cell-cell junctions. The lamina propria is a connective tissue that underlies these cells

Question 4

Types of cell-cell junctions (5)

Answer 4

1. Adherens – cadherin family
2. Desmosomes – cadherin family
3. Tight junctions – claudins & occludins
4. Gap junctions – connexins, innexins
5. Transient (selective) junctions – selectins, Ig superfamily, & integrins

Question 5

Functions of cell-cell junctions (2)

Answer 5

1. Bring neighboring cells together
2. Anchor the cytoskeletons of each cell

Question 6

Cadherins

Answer 6

Make up adherens junctions- they depend on calcium for binding. There are 2 categories- classical and nonclassical. Both categories act similarly in a symmetrical and homophilic reaction. They have contact with the actin part of the cytoskeleton. The actin of one cell is connected to a cadherin, which interacts with a cadherin of a neighboring cell, which in turn is interacting with that cell’s actin cytoskeleton. The intracellular part of the cadherin goes through the membrane and into the cytoplasm . The intracellular part is indirectly linked to the actin portion of the cytoskeleton. This is mediated by catenin

Question 7

Classical cadherins

Answer 7

Closely related in sequence. Serve adhesive & signaling functions

Question 8

Nonclassical cadherins

Answer 8

Distantly related in sequence. Some adhesive – desmocollins, desmogleins (desmosomes)
Some primarily involved in signaling

Question 9

Cadherin binding

Answer 9

Cadherins bind to one another at their N-termini
Each cadherin has a terminal knob that fits into the binding pocket of the other cadherin (binding is symmetrical). Cadherin-cadherin binding is of low affinity, but having a lot of the protein (high avidity) makes up for this

Question 10

Adherens structure

Answer 10

Consists of many repeats of cadherin. The number of repeats varies, but there are usually 5 repeats for classical cadherins. These junctions are linked to the actin cytoskeleton. As they pull the cells together, the junctions form an adhesion belt (cadherins) close beneath apical face of epithelium

Question 11

Cadherin structure

Answer 11

The extracellular portion of cadherins is repeats of cadherin proteins. In between the repeats, there are hinge regions connecting the repeats (they are called hinges because they are flexible). Calcium binds to sites near the hinge to prevent flexing and add rigidity to the cadherin

Question 12

β-catenin

Answer 12

Found in classical cadherins, located at the tip of the cytoplasmic end of the cadherin. It couples cadherin to actin via other anchor proteins (catenins), which in turn is linked to the actin cytoskeleton

Question 13

P120-catenin

Answer 13

Found in classical cadherins, helps regulate assembly of β-catenin with the other anchor proteins

Question 14

Adhesion belt

Answer 14

Found close beneath apical face of the epithelium. It links the actin bundles/actin cytoskeletons of adjacent cells. It creates a transcellular network, allowing the cells to behave together, which contracts and rearranges the epithelium in a unified manner

Question 15

Desmosomes

Answer 15

A type of cell junction linking the intermediate filaments of the cytoskeleton- it links the intermediate filaments of adjacent cells. The intermediate filaments are bound to desmosomes all over cell on the lateral sides & to hemidesmosomes at basal surface. This creates a lattice-like network that gives epithelial tissue great mechanical strength

Question 16

Tight junctions functions (3)

Answer 16

1. Prevent leakiness of the epithelium
2. Membrane domain “fences”
3. Paracellular transport

Question 17

Tight junctions

Answer 17

Contribute to the main function of epithelial tissue, which is to be a barrier to the outside world. It prevents anything from leaking in and prevents transcytosed material from leaking out. It also acts as a barrier and keeps the apical membrane proteins from mixing into the lateral and basal membranes. Tight junction proteins include the claudins, occludins, and tricellulin

Question 18

Paracellular transport

Answer 18

When tight junctions are transiently altered to permit the flow of solutes through it. This is important for the absorption of amino acids and monosaccharides from intestinal lumen

Question 19

Claudins

Answer 19

Main transmembrane proteins that form the sealing strands of the tight junctions. They can also form paracellular pores and allow for cell-cell interactions

Question 20

Occludins

Answer 20

Not as essential as claudins, but they play a role in forming tight junctions

Question 21

Tricellulin

Answer 21

Required to seal membranes together, prevent transepithelial leakage at points where 3 cells meet

Question 22

Focal connections

Answer 22

Tight junctions form focal connections- many of the claudins and occludins making up these junctions are interacting with each other. Any time neighboring cells come together, it is a focal connection of tight junctions

Question 23

Gap junctions

Answer 23

The formation of many intercellular channels between neighboring cells which allows for exchange of molecules, like a molecular sieve. Formed by proteins called connexins and innexins. Allows for inorganic ions and other water soluble molecules to pass between cells- the neighboring cells can be coupled metabolically and electrically. The largest pore a gap junction can form is 1.5 nanometers in diameter, which is big enough for inorganic ions, sugars, amino acids, nucleotides, vitamins, and cAMP. However, it is too small for macromolecules (proteins, nucleic acids, and polysaccharides)

Question 24

Connexins

Answer 24

4-pass transmembrane proteins. 6 of them come together in quaternary structure to form a hemichannel (half channel, referred to as a connexon). 2 connexons from adjacent cells come together to form intercellular channels, which are part of the gap junctions

Question 25

Functions of gap junctions (3)

Answer 25

1. Rapid cell-cell spread of action potentials in neurons
2. Couples contraction of cardiac and smooth myocytes
3. Sharing of metabolites among epithelial cells

Question 26

When are gap junctions found in neurons?

Answer 26

In electrical synapses, which couple together the presynaptic and postsynaptic neurons so there’s no need for neurotransmitters to travel across the synapse. Occurs in situations where parallel neurons are firing in synchrony. They are essential for the function of the nervous system

Question 27

Gap junction permeability

Answer 27

The channels can exist in open or closed states. Their permeability can be reduced by changes in pH and increases in cytoplasmic calcium. Increase in calcium is usually considered a danger signal because calcium may be interpreted as a sign of membrane rupture or cell death. Permeability decrease prevents the spread of the toxic signal to healthy neighboring cells

Question 28

Selectins

Answer 28

Transient cell-cell adhesion proteins that require calcium. They are expressed on the endothelial cells of blood vessel walls during inflammation. It helps to slow down neutrophils and other cells that are part of the inflammatory response. Selectins capture neutrophils by binding to sugars on the neutrophils (therefore, they are selective lectins)

Question 29

E-selectins

Answer 29

The selectins found on endothelial cells- they bind to sugars on leukocytes, like neutrophils. The first binding interaction causes a weak adhesion where the inflammatory cells continue to move, but they roll slowly. Then, there is a second binding event where integrins on the leukocytes are activated and interact with other molecules in the endothelial cells (I-CAMs, V-CAMs from the immunoglobulin superfamily). This results in strong adhesion- the inflammatory cells are immobilized and then go through the process of diapedesis

Question 30

Diapedesis

Answer 30

Leukocyte migration out of the blood vessels and into the underlying tissue

Question 30

Ig superfamily

Answer 30

Includes intercellular cell adhesion molecules (ICAMs) and vascular cell adhesion molecules (VCAMs). They interact with integrins on the inflammatory cell surface, which allows for the tight adhesion in diapedesis. All have Ig-like domains with disulfide bonds

Question 31

Selectins and diapedesis (2 steps)

Answer 31

1. Weak adhesion and rolling is selectin dependent
2. Strong adhesion and emigration is integrin dependent- interacts with integrins on the leukocyte surface

Question 32

Components of the ECM (2)

Answer 32

1. Epithelial tissue- connected on the basal side to a type of ECM called the basal lamina
2. Connective tissue- made of lots of ECM with sparsely distributed cells

Question 33

Which cells synthesize the ECM?

Answer 33

It is synthesized by cells on each side of it

Question 34

2 main classes of macromolecules making up the ECM

Answer 34

1. Fibrous glycoproteins (short oligosaccharide chains)
2. Proteoglycans- modified by glycosaminoglycans (GAGs) linked to core proteins. This makes up the largest proportion of the ECM

Question 35

Fibrous proteins in the ECM

Answer 35

Collagen, fibronectin, and other fibrous proteins contribute. They form a meshwork in the basal lamina

Question 36

Basal lamina

Answer 36

A thin sheet of ECM found at the basal end of epithelial cells. It is also called the basement membrane, and is 40-120 nanometers thick. In addition to epithelial cells, the basal lamina also wraps around myocytes, adipose cells, and Schwann cells of the myelin sheath. Its functions include mechanical connection of cells to underlying connective tissue, and plays a mechanical role in anchoring the epithelium. The epidermis is one example, it is an epithelial covering

Question 37

Junctional epidermolysis bullosa

Answer 37

Genetic defect in basal lamina proteins don’t anchor the epidermis properly, which results in blistering. This demonstrates the importance of the basal lamina in anchoring the epithelium

Question 38

Laminin structure

Answer 38

Flexible proteins with 3 long polypeptide chains (α, β, g) in a quaternary structure, held together by S-S disulfide bonds. Its structure is referred to as an asymmetric bouquet. β and g heads promote self assembly of multiple laminins into a meshwork. This forms first and lays down a meshwork for the basal lamina. α heads bind integrins on cells

Question 39

Collagen structure

Answer 39

A fibrous glycoprotein with a triple stranded helix. The helix is made of 3 collagen polypeptides (α chains) wound around 1 another in a quaternary structure, forming a ropelike superhelix. Collagen is rich in proline and glycine. Pro ring stabilizes helix, Glycine (every 3rd residue) provides flexibility for helix formation

Question 40

Type IV collagen

Answer 40

A key component of the basal lamina. Interrupted in >20 regions = flexibility

Question 41

Which cells synthesize the ECM of connective tissue?

Answer 41

Produced by cells in the matrix- Fibroblasts, osteoblasts, chondroblasts

Question 42

Components of the ECM of connective tissue (2)

Answer 42

1. Proteoglycans with GAGs
2. Fibrous glycoproteins- like collagen (adds strength) & elastin (makes it resilient)

Question 43

Ground substance

Answer 43

Formed when proteoglycans and fibrous glycoproteins come together. It is a highly hydrated, gel-like substance formed mainly by proteoglycans. Fibrous proteins are embedded are add strength

Question 44

Glycosaminoglycans (GAGs)

Answer 44

Unbranched (linear in size and shape) polysaccharide chains of repeating disaccharides- 1 is always an amino sugar, and the other is usually uronic acid sugar (contains carboxylic acid). GAGs are different from other sugars in that they are heavily sulfated and therefore have a negative charge. They make up the bulk of the ECM

Question 45

4 subgroups of GAGs

Answer 45

The 4 groups are distinguished by sugars, linkages & sulfate groups
1. Hyaluronan
2. Chondroitin sulfate/dermatan sulfate
3. Heparin sulfate
4. Keratin sulfate

Question 46

GAGs in the ECM

Answer 46

GAG molecular weight takes up the bulk of the area, but it makes up less than 10% of the weight of fibrous proteins in connective tissue. This is due to the unbranched structure of GAGs, which allows it to fill up a lot of space. The negative charge of GAGs attracts a cloud of cations, especially sodium. This causes osmotic action, which sucks up large amounts of water into the ECM. Creates turgor pressure that enables ECM to resist compressive forces. Example- cartilage ECM of the knee supports loads of pressure due to this

Question 47

How do GAGs fill up the ECM?

Answer 47

One molecule of hyaluronan (a GAG) can easily fill the ECM space, despite its low molecular weight. It is able to fill the ECM due to its size and winding shape.

Question 48

Proteoglycans

Answer 48

When GAGs are covalently bonded to proteins- this includes all GAGs except hyaluronan, which does does not attach to proteins. Proteoglycans are created through O-linked glycosylation as proteins move through the Golgi. Enzymes called glycosyl transferases add GAG sugars 1 at a time, so a large polysaccharide can be part of the protein structure. A proteoglycan must at least have one GAG connected to it by definition. Proteoglycans are 95% carbohydrate by weight and are usually 80 sugars long. This is in contrast to glycoproteins, which are only 1-60% carbohydrate by weight

Question 49

O-linked glycosylation

Answer 49

Occurs when sugars are being added to proteins as they move through the Golgi. It is called O linked because sugars are added to the hydroxyl group of the amino acids serine or threonine. The sugars here are added one at a time, like an extension of N-linked oligosaccharides. When a protein undergoes O-linked glycosylation, that usually means that it is heavily glycosylated. This type of glycosylation is heaviest on mucins and proteoglycan core proteins.

Question 50

Type 1 collagen

Answer 50

A fibrous glycoprotein that is part of the ECM of connective tissue. It has a long, stiff, triple-stranded helix. Less flexible than it is in the basal lamina, and more stiff than it is in connective tissue. 3 collagen polypeptides (α chains) are wound around 1 another to form a ropelike superhelix. Collagen is rich in Pro & Glycine. The proline ring stabilizes helix, and Glycine (every 3rd residue) provides flexibility for helix formation

Question 51

Elastic fibers

Answer 51

Another type of fibrous protein found in the ECM of connective tissue. Elastin is their main component. Elastic fibers are interwoven with collagen fibrils, which prevents overstretching and tearing and allows the tissue to be resilient. They are rich in proline and glycine, but usually are not glycosylated. Part of the polypeptide of elastin adopts a loose “random coil”, which allows the fibers to stretch and recoil. Elastic fibers are dominant in the ECM of arteries. They stretch with the smooth muscle and prevent arterial tension

Question 52

Elastin random coil

Answer 52

The elastin forms a random coil in the relaxed form, but can easily be stretched out without breaking. It can also bounce back to its original form after stretching. This adds resiliency to the ECM

Question 53

Integrins

Answer 53

Cells in the ECM are able to interact with the ECM through integrins, as integrins bind to ECM proteins (matrix receptors). They transmit signals from ECM to cell & vice versa. Tension causes integrins to tighten grip on what they’re holding on to in the cytoplasm and what they’re holding onto in the ECM, while loss of tension does the reverse. Responsible for leukocyte adhesion to endothelial cells

Question 54

Integrin structure

Answer 54

Contains 2 glycoprotein subunits – α & β. It has a large N-terminus extracellular domain that binds to and interacts with ECM proteins. Integrins traverse the membrane and have short intracellular tails in the cytoplasm. The β subunit tail binds to a binding protein called talin.

Question 55

Talin

Answer 55

A binding protein that binds to the β subunit of integrins. Talin links integrins (& ECM) to the actin cytoskeleton. Talin is essential for the ability of the integrins to link the actin cytoskeleton to the ECM. Actin-ECM integrin junctions may be small, transient or large, durable

Question 56

Hemidesmosomes

Answer 56

Integrin-dependent linkages of epithelial cells to the basal lamina. They link to keratin of the intermediate filaments and to laminin in the basal lamina.
Plectin & dystonin serve as adaptor proteins. The keratin of the epithelial cells is linked to the integrin through these adaptor proteins. Integrin binds to the adaptor proteins and from there, binds to collagen and laminin. In essence, the adaptor proteins allow integrin to link the epithelial cells to the basal lamina beneath them

Question 57

Outside-in activation of integrins (5)

Answer 57

1. In their inactive state, integrins are folded, α-β domains are closely associated.
2. Integrins are activated to bind when the ECM ligand is present, and the extracellular domain unfolds. This stimulus is coming from the outside
3. Transmembrane and intracellular contacts are broken
4. Tails move apart, the talin binding site on the β subunit is exposed
5. Assembly of actin anchored to intracellular end of integrin

Question 58

Inside-out activation (4 steps)

Answer 58

1. When integrins are activated from inside the cell. Talin competes w/ α for β chain binding
2. Talin binding undoes intracellular α-β linkage, the extracellular binding site is exposed so it can bind to ECM proteins
3. The PI(4,5)P2 lipid binds/activates talin to induce binding to β domain
4. PI(4,5)P2 is produced in response to G-proteins & receptor Y kinases. It links extracellular signals to integrin activation