Structure and Function of ECM

Question 1

Basement Membrane organization

Answer 1

1. Basement membrane
   
   1. Basal Lamina
      
      1. Lamina rara/lucuda/interna: Lamin 5, Collagen 17
      2. Lamina densa: Lamin 1, Collagen 4, Perlecan
   2. Lamina reticularis: Type 3 collagen –> made by fibroblast

Question 2

Components of ECM

Answer 2

1. Collagen
   
   1. Impart strength
   2. Structural protein
2. Elastin
   
   1. Impart resilience
   2. Structural protein
3. GAGs
   
   1. Combine with water to resit compression
4. Proteoglycans
   
   1. Proteins covalently bound to GAG chain
5. Adhesive glycoproteins
   
   1. Fibronectin, Laminin, Entactin

Question 3

Collagen Stucture

Answer 3

1. Individual collagen chains are made with a distinct repeating amino acid sequence of Gly-X-Y
   
   1. Gly has H side chain that is wrapped in middle of helix
   2. X and Y can be prolines and lysines that are hydroxylated so that hydrogen bonding can occur within monomer
2. Individual collagen chains can assemble into a triple helix
3. Fibril forming collagens can form fibrils from triple helix collagens
4. ***A single mutation in amino acid of collagen can destablilize it and lead to disease state

Question 4

Fibril forming collagens

Answer 4

Collagens I, II, III, V, XI

Question 5

Fibril associated Collagens

Answer 5

Collagens IX and XII

Question 6

Network Forming Collagens

Answer 6

Collagens IV and VII

Question 7

Transmembrane Collagens

Answer 7

Collagens XVII

Question 8

Specific disease states associated with different collagen types

Answer 8

Collagen I: in bone

Collagen II: in cartilage

Collagen III: in skin and blood vessels

*All are fibril forming

Question 9

Biosynthesis of Fibril Forming Collagens

Answer 9

1. Pro-alpha collagen chain is secreted into the ER/Golgli
2. Pro-alpha chain is hydroxylated at proline and lysine residues
3. Pro-alpha chain is glycosylated at lysine residues
4. Pro-alpha chain assembles into a procollagen triple-helix
5. The pro-collagen is secreted from Golgi out of the cell
6. The pro-collagen extension peptides at the N and C-terminus are cleaved to make tropocollagen
7. Tropocollagen can self assemble into a fibril

• Pro-alpha collagen –> pro-collagen –> tropocollagen –> fibril
• Biosynthesis of collagen occurs inside and outside of cell
• Type I,II, III, V, and XI can form collagen fibrils

Question 10

Fibril associated collagen

Answer 10

1. Fibril associated collagens form a hinge on the fibril forming collagens and cause a interruption of the triple helix such that fibril forming collagens are less rigid
2. Fibril associated collagens still have extension peptides are associated
3. Type IX and Type XII
   
   1. Mutation in Type IX collagen lead to epiphyseal dysplasia such that you become arthritic

Question 11

Network forming collagen

Answer 11

1. Network forming collagens don’t have their extension peptides cleaved at N and C-terminus
2. Extension peptides interact to form chicken wire array
   
   1. C-terminus heads dimerize
   2. N-terminus tails project above and below plane to form sheetlike meshwork
3. Type IV and Type VII collagens
   
   1. Type VII is a network forming collagen that keeps stroma attached to lamina densa.
   2. Mutations in this lead to blistering disease in which epithalial cells peal away from lamina densa

Question 12

Transmembrane collagen

Answer 12

1. Helps bind epithelial cells to lamina densa
2. Type XVII collagen
   
   1. Mutation causes epithelial cells to peel away causing blistering disease.

Question 13

Elastin

Answer 13

1. Elastin crosslinks with each other via unique amino acids forming rubber bands
2. Assembled extracellularly like collagen
3. Provides resilience and flexibility to tissue
   
   1. Aorta
   2. Blood Vessels
   3. Heart- Type III and elastin fibers
4. Elastins are made from fibrils
5. Mutation in fibrillin 1
   
   1. Marfan syndrome
      
      1. Autosomal dominant disorder
      2. Recoil that keeps limbs in proportion does not occur
      3. Aorta prone to rupture

Question 14

Glycosamine Glycans (GAGs) and Examples

Answer 14

1. GAGs are long sugar molecules made of repeating disaccharides
2. At physiological pH, carboxyl group are negative:
   1. These molecules repel each other and they extend away from each other taking up a lot of space
   2. They attract water and are major components of synovial fluid in joints and vitreous humor in the eye
   3. I.e. Hyaluronic acid
      
      1. Long
      2. Not sulfated
   4. Chondoitin sulfate and Heparan Sulfate
      
      1. Sulfated and can attach to protein

Question 15

Proteoglycans

Answer 15

1. Proteins + GAGs
   
   1. Aggrecan
      
      1. Location: cartilage
      2. GAGs: Chondroitin sulfate and keratin sulfate
      3. Binds type II and type IX collagen
      4. Function: mechanical support
   2. Perlecan
      
      1. Location: basal lamina
      2. GAGs: heparan sulfate
      3. Function: structural and filtering function in basal lamina
   3. Syndecan-1
      
      1. Location: fibroblast and epithelial cell surface
      2. GAGs: chondroitin sulfate and heparan sulfate
      3. Function: cell adhesion and is a co-receptor for FGF
         
         1. FGF does not form a dimer
         2. Syndecan uses heparin sulfate branches to bind FGF such that receptor dimerizes.

Question 16

Adhesive Glycoproteins

Answer 16

1. Have different modules along length that can interact with different extracellular molecules
2. Therefore, you can form networks
3. These are molecules that cells can attach to in the ECM
4. They have specific sequences (RGD amino acid sequence) that can interact with specific receptors (integrins) in the cell surface

Question 17

Laminin

Answer 17

1. Structure
   
   1. Triple Helix
   2. Has cross-like configuration
2. Has different regions that can bind different molecules in ECM
   
   1. Cell Binding domain
   2. Collagen binding domain
   3. Proteoglycan heparans sulfate binding domain
   4. RGD sequence to bind integrins

Question 18

Fibronectin

Answer 18

1. Structure
   
   1. Dimer held together by disulfide bonds
2. Has different regions that can bind different molecules in ECM
   
   1. Collagen binding domain
   2. Proteoglycan binding domain
   3. Clotting factors like fibrin binding domain
   4. Has RGD sequence that binds to integrins.

Question 19

Integrins

Answer 19

1. Exist as alpha/beta dimers
   
   1. Only beta subunit interacts with actin cytoskeleton
      
      1. Exception: Integrin a6b4 in hemidesmosomes interacts with intermediate filaments
   2. Activation of integrins
      
      1. Integrins cluster after binding substrate
      2. Integrins then bind actin-binding proteins
      3. Actin binding proteins bind actin
      4. Focal adhesion forms at site of bound integrin via Rho GTPase
      5. Focal Adhesion Kinase is recruited
      6. FAK autophosphorylates tyrosine residues on itself and activates downstream molecules.

Question 20

Different types of Integrins

Answer 20

1. Beta 1: binds with fibrinectin and laminin
2. Beta 2: found on WBC and bind to endothelial surface
3. Beta 3: bind to fibrinogen on platelets and form clots
4. Beta 4: found in hemidesmosomes and bind specific laminins

Question 21

Hemidesmosomes

Answer 21

1. Bind to Lamin V
2. Bind to Type XVII collagen
   
   1. Both are found in lamina rara
   2. Mutations in either can cause blistering disease-Bullosa Pemphigoid.