ECM and integrin receptors

Question 1

Describe the ECM and its roles.

Answer 1

• complex meshwork of extracellular molecules
• secreted and assembled by local cells (fibroblasts)
• provides supportive framework to hold cells and tissues together
• found in multicellular organisms only
• critical for development of distinct tissues and organs
• regulates many cellular processes through interaction with integrins

Question 2

Describe integrin receptors and their roles

Answer 2

• they are the principle matrix receptor in animal cells
• membrane-spanning receptors that provide a link between the inside of the cell and the ECM
• bind to ECM proteins to activate intracellular signaling pathways
• transduce cells bidirectionally across the membrane (inside out and outside in)
• only in multicellular organisms
• regulates many cellular behaviors

Question 3

Describe the role of ECM and integrins in cancer.

Answer 3

Changes in ECM and integrin expression are associated with many pathological conditions, including cancer.

• ECM serves as barrier to tumor progression (basal lamina)
• alterations in ECM (tension) can contribute to tumor progression
• ECM and integrins can be exploited to target tumors therapeutically

Question 4

Which transmembrane adhesion protein is responsible for transducing the activating signal to focal contact-associated kinase (FAK)?

Answer 4

integrin

Question 5

What are the components of the ECM?

Answer 5

• GAGs: unbranched polysaccharide chains composed of repeating disaccharide units usually attached to a core protein. When attached to the protein it is called a proteoglycan
• fibrous proteins: collagen, elastic fibers, fibronectin

Question 6

What are the four types of GAGs?

Answer 6

1. hylauronan (simplest GAG)
2. heparan sulfate
3. chondroitin/dermatan sulfate (important in joints)
4. keratin sulfate

Question 7

How do different types of GAGs differ?

Answer 7

differ based on types of sugars, linkage between sugars, and sulfation of sugars

Question 8

Describe complex GAGs.

Answer 8

they are sulfated to add a negative charge, thus attracting sodium ions and H2O to keep the ECM well hydrated and at a large volume. One example of this type of GAG is heparan sulfate.

Question 9

Describe the proteoglycan core protein.

Answer 9

• core protein binds GAG molecules. One bound is a decorin
• many bound is an aggrecan
• core proteins are glycosylated and secreted through ER-golgi-mediated secretory pathway

Question 10

What are the benefits of the large size of GAGs?

Answer 10

Because the size is due to hydration and sweeling, they enable the matrix to withstand compressive forces while allowing rapid diffusion of nutrients (not too dense) between blood and tissue cells. Hyaluronan allows tissues to resist compression and are found in ECM of load-bearing joints.

Question 11

What are the functions of proteoglycans and GAGs?

Answer 11

• selective sieve to regulate molecular trafficking
• regulation of chemical signaling (e.g. heparan sulfate can bind and bring FGF to FGFR)
• regulation of other secreted proteins
• act as co-receptors

Question 12

What are some examples of GAGs and proteoglycans regulating other secreted proteins?

Answer 12

heparan sulfate can immobilize and sequester chemokines together for an immune cell;

=dermatan sulfate can organize collagen;

• others can sterically block protein activity;
• others can protect proteins from proteases

Question 13

How do proteoglycans and GAGs act as co-receptors?

Answer 13

• some proteoglycans are tethered to the plasma membrane of cells, and therefore aid in the activation of cellular receptors
• syndecans: heparan sulfate present FGF to FGFR
• betaglycans: present TGF-beta to TGF-beta receptor

Question 14

What is Hurler’s syndrome?

Answer 14

• autosomal recessive
• accumulation of heparan sulfate and dermatan sulfate as a result in deficiency in the enzyme which degrades them
• cells cannot grow normally because ECM is overgrown
• causes dwarfism, mental retardation

Question 15

Role of collagen in ECM?

Answer 15

provides tensile strength to matrix. Found in all multicellular organisms.

Question 16

Describe the structure of collagen.

Answer 16

• 3 alpha helices wrapped around each other
• glycine allows for close interaction of 3 helices
• proline rings stabilize the helices
• hydroxyproline, hydroxylysine residues also stabilize the triple helix

Question 17

How many known types of collagen are there?

Answer 17

40

Question 18

Describe the steps of fibrillar collagen formation.

Answer 18

1. pro-alpha chain is made in the ER/golgi
2. selected prolines and lysines are hydroxylated
3. selected hydroxylysines are glycosylated
4. three pro-alpha chains are self-assembled
5. procollagen triple helix is formed and leaves golgi in secretory vesicle
6. procollagen is secreted from the cell
7. propeptides are cleaved (N and C termini) to prevent collagen aggregation
8. collagen molecules self-assemble into fibrils, directed by crosslinking of lysine residues
9. collagen fibrils aggregate to form a collagen fiber

Question 19

How do cells determine size/organization of collagen fibrils?

Answer 19

• alter collagen expression by changes in gene expression, and by protein turnover (degradation)
• assembly is guided by invagination of the plasma membrane
• organization into different types/patterns of collagen is aided by secretion of collagen-interacting proteins
• tension

Question 20

What are the different patterns of collagen organization?

Answer 20

• wickerwork pattern: collagen fibers are oriented perpendicularly to each other (e.g. skin)
• bundled pattern: collagen fibers are oriented parallel to each other (e.g. tendon). these are the stronger type

Question 21

How are collagen fibrils linked together?

Answer 21

Fibril-associated collagens link fibrils to each other and to other ECM components. Also determine nature of collagen fibrils.

Question 22

Describe fibril-associated collagens.

Answer 22

• type IX (cornea) and type XII (tendon) collagens
• triple helix structure is interrupted with nonhelical segments to allow for bending
• retains propeptides after secretion and don’t form fibrils
• bind to surface of fibrillar collagens and to other ECM components to determine organization of collagen matrix

Question 23

Describe how tension helps determine organization of collagen fibrils.

Answer 23

cells tug on collagen, reorganize it, and use it to migrate out. This can be seen in heart fibroblasts, where collagen fibers form between two explants.

(ECM can also influence the behavior of the cells it supports. Not just one direction!)

Question 24

Collagen-associated diseases?

Answer 24

• scurvy: vitamin C deficiency. vit C acts as cofactor for hydroxylation of lysines and prolines
• osteogenesis imperfecta: collagen type I defect (glycine substitution). brittle bones, sclera of eye is blue
• scleroderma: autoimmune disease causes buildup of collagen in skin and organs (skin tightens and fingers lose mobility)

Question 25

How is the ECM able to recoil after transient stretch?

Answer 25

Elastic fibers allow for recoiling. They can recoil after stretching 1.5 times their length. This is very important in skin, blood vessels, and lungs. The network of elastic fibers interwoven with nonelastic collagen fibrils regul\ates the elasticity of tissues.

Question 26

What are elastic fibers composed of?

Answer 26

- elastin is primary component - rich in lysine and proline (some hydroxyproline) - tropoelastin molecules are secreted from cell where they assemble in association with plasma membrane and generate fibers from lysine crosslinking - random coiled assembly allows stretching and retratction (hydrophobic regions for elasticity, and alpha helices for crosslinking) - deficiency leads to artery narrowing

Question 27

How is elastin deposition directed?

Answer 27

early in development, microfibrils are secreted into ECM and provide scaffolding for elastin. later in development microfibrils help organize elastin into bundles Microfibrils are composed of glycoproteins such as fibrillin mutation in fibrilin gene causes Marfan's syndrome

Question 28

What is fibronectin?

Answer 28

- large glycoprotein important in cell-ECM interactions - consists of a dimer with 2 large polypeptide chains joined by disulfide bond at the C termini - each subunit contains 5-6 functionally distinc domains for binding to a specific molecule or cell (e.g. cell-binding domain, collagen-binding domain, heparan-binding domain)

Question 29

What mediates fibronectin binding to integrin?

Answer 29

The type III FN repeat contains an RGD residue sequence which binds to integrins. Synergy sequences at this domain can also bind integrin.

Question 30

How is fibronectin fibril formation mediated?

Answer 30

fibronectin normally travels throughout the blood, and only forms fibrils on surface of cells, which requires integrins: 1. fibronectin dimers bind integrins 2. linkage to intracellular cytoskeleton causes tension which pulls fibronectin into an open conformation, revealing cryptic binding sites 3. fibrils can then self-assemble

Question 31

describe the basal lamina.

Answer 31

- thin, flexible sheet of ECM that underlies all epithelial cells. - also *surrounds* muscle (rather than just on one side of the cell) and schwann cells - provides support and regulatory signals to cells it encases.

Question 32

components of the basal lamina?

Answer 32

- laminin: organizes basal lamina sheet - type IV collagen: provides tensile strength - nidogen and perlecan: links laminins and collagen

Question 33

Describe laminin.

Answer 33

- primary component of basal lamina - heterotrimeric protein assembled in shape of assymetric cross. (three heads + tail region) - combinations of chain types can yield different types of laminins - contain binding sites for other laminins and integrins to make a laminin network - loss of laminin-gamma-1 is embryonic lethal

Question 34

How is basal lamina assembled?

Answer 34

1. components are secreted from cells 2. tails anchored by matrix receptors, allowing heads to interact 3. laminin coordinates assembly of laminin and collagen networks by nidogen and perlecan linkers

Question 35

What is epidermolysis bullosa?

Answer 35

genetic defect in basement membrane resulting in severe skin blistering

Question 36

What are the functions of the basal lamina?

Answer 36

1. provides cells with mechanical support (epidermolysis bullosa defect) 2. compartmentalization of tissues (protects cells from harsh environments) 3. selective filter (e.g. kidneys separate blood from urine) 4. scaffold during tissue regeneration (e.g. neuromuscular junction) 5. act in concert with integrins to regulate cell polarity, metabolism, survival, proliferation, and differentiation

Question 37

What is the purpose of protein turnover in the ECM?

Answer 37

Degradation of the ECM is necessary for tissue repair, remodeling, adaptation to stress, proliferation, and migration -also a requirement for cancer metastasis

Question 38

What protease degrades the ECM for remodeling in pregnancy?

Answer 38

MMP proteases (matrix metalloprotease)

Question 39

Name the two classes of extracellular proteases.

Answer 39

1. matrix metalloproteases (MMPs): depend on calcium or zinc 2. serine proteases: reactive serine in active site

Question 40

Describe the regulation of ECM protease activity.

Answer 40

1. local activation: proteases are secreted in inactive form and activated through cleavage only in areas where they are needed 2. confinement by cell-surface receptors 3. secretion of protease inhibitors 4. protease specificity (e.g. collagenase only cleaves collagen)

Question 41

Different types of ECM protease inhibitors?

Answer 41

- tissue inhibitors of MMPs (TIMPs) - serpins inhibit serine proteases