EMC and adhesion - exam Flashcards
IMPORTANCE
Example to illustrate importance:
- mouse mammary gland, an epithelial tissue
IMPORTANCE
Cell-cell adhesion and cell to EMC adhesion are essential to normal tissue structure and function
WHAT IS EXTRACELLULAR MATRIX
- the complex network of secreted proteins and carbohydrates that fills the space between cells
- at most basic level it fills space in-between cells
- function aspect:
o connective tissue vs the basal lamina
WHAT IS EXTRACELLULAR MATRIX
organisation of the basal lamina
The basal lamina is a specialized sheet of extracellular matrix (ECM) found at the base of epithelial, muscle, and nerve cells. It’s secreted by the cells it supports and is made mainly of:
Laminin (key organizer)
Type IV collagen (forms a supportive mesh)
Nidogen/entactin (linking proteins)
Perlecan (a heparan sulfate proteoglycan)
WHAT IS EXTRACELLULAR MATRIX
The basal lamina in different tissues
muscle
epithelium
kindney glomerulus
key functions of basal lamnia
Structural support
Cell anchoring
Barrier/filter (especially in the kidney)
Tissue organization and repair
Signaling (via integrins and other receptors)
WHAT IS EXTRACELLULAR MATRIX
The basal lamina in different tissues
- epthelium
Location: Lies beneath epithelial cells, separating them from underlying connective tissue.
Function:
Provides physical support.
Acts as a selective barrier to regulate exchange between epithelial cells and stroma.
Guides cell polarity and differentiation.
Structure:
Often has two layers when viewed with EM:
Basal lamina (cell-derived ECM layer).
Reticular lamina (deeper collagen-rich layer from connective tissue).
WHAT IS EXTRACELLULAR MATRIX
The basal lamina in different tissues
- muscle
Location: Surrounds individual muscle fibers (myofibers).
Function:
Provides a scaffold during muscle regeneration.
Anchors muscle cells via dystroglycan–laminin–integrin complexes.
Important for force transmission from inside the fiber to connective tissue.
Clinical note: In diseases like Duchenne muscular dystrophy, defects in linking proteins (like dystrophin) impair connection to the basal lamina.
WHAT IS EXTRACELLULAR MATRIX
The basal lamina in different tissues
- kidney glomerulus
Location: Forms part of the glomerular filtration barrier, between the endothelium and podocytes.
Function:
Acts as a highly selective molecular filter, allowing water and small solutes to pass while blocking proteins.
Critical for urine formation.
Structure:
Thicker and denser than in other tissues.
Made by both endothelial cells and podocytes.
Clinical note: Damage or mutations in type IV collagen (e.g., Alport syndrome) lead to proteinuria and kidney dysfunction.
the extracellular matric is made and organised by the…
cells within it
in connective tissues the macromolecules that make up the extracellular matrix are…
secreted by cells called fibroblasts
the basal lamina mainly synthesized by..
the cells that rest on it
e.g. basal keratinocytes of the epidermis
Simple experiment to illustrate the importance of extracellular matrix
Dissect alveoli from pregnant mice
* Disaggregate cells
* Plate out in culture (in lab)
* Give choice of two different extracellular proteins
o Laminin
o Fibronectin (not found in basal lamina - embryogenesis and wound healing)
function of ECM
- Space filler (e.g. hyaluronan)
- Tissue organizer (e.g. laminin)
- Protective
- Cell migration
- Regulation of cell function (e.g. laminin)
ECM as tissue organiser
- When interact with laminin self organism into spherical structures
o Not dissimilar to the alveoli - No difference apart from protein
- Must be a signal from interacting with laminin that causes it to organise this way
ECM as a regulator of cell function
- In this experiment monitored milk production
fibronectin + prolactin
–> b-casein not produced
laminin + prolactin
–> b-casin produced and secreted into the interior of spherical structures
COMPONENTS OF THE ECM
- Fibrous proteins
a) Structural (e.g. collagens)
b) Adhesive (e.g. laminin, fibronectin, collagens) - Glycosaminoglycans
o E.g. hyaluronan - Proteoglycans
o E.g. aggrecan
COMPONENTS OF THE ECM
fibrous proteins
- Collagens: have structural and adhesive role
o Insoluble, extracellular glycoproteins
o Most abundant protein in the human body
25% of protein mass
o Essential structural component of all connective tissues including: bones, tendons, cartilage, and skin
o Also an important component of the basal lamina
Lattice grid on which everything else is assembled
COMPONENTS OF THE ECM
fibrous proteins
- assembly of collagen
into collagen fibres
o Fibrils organised into collagen fibre
Highly ordered strength giving molecules in connective tissue
o Osteogenesis imperfecta (‘brittle bone disease’)
Caused by a mutation in the gene for collagen type I
* COL1A2
COMPONENTS OF THE ECM
GAGs – Glycosaminoglycans
- Unbranched polysaccharide chains composed of repeating disaccharide units
o Highly negatively charged - GAGs are inflexible and adopt highly extended structures that occupy a large volume reflective to their mass and form gels at even quite low concentrations
- The high density of negative charges attracts a ‘cloud’ of cations (e.g. Na+) causing large amount of water to be sucked into the matrix by osmosis
- Good for resisting compressive forces
o In contrast to collagen, which resists stretching forces
COMPONENTS OF THE ECM
GAGs – Glycosaminoglycans
Example of a GAG: hyaluronan
- Up to 25,000 repeating disaccharide units
- Specialised example
o Space filler
Can occupy a lot of space
COMPONENTS OF THE ECM
GAGs – Glycosaminoglycans
hyaluronan not a typical GAG
- All other GAGs
o Have different disaccharide subunits arranged in a more complex manner
o Have much shorter chains (<300 sugars)
o Are covalently linked to protein to form proteoglycans
COMPONENTS OF THE ECM
Structure of a typical proteoglycan
- E.g. aggrecan (protective function)
o Aggrecan forms aggregates which are important in cartilage function
Provides a protective layer and lubricating surface
o Very large macromolecule
o Can see it under electron microscope quite easily
COMPONENTS OF THE ECM
Adhesive glycoproteins
- E.g. fibronectin, laminin, fibrinogen, vitronectin, and collagen
- With the exception of collagen, they are relatively minor components, but their importance is illustrated by knockout mice experiments
- Laminin alpha2 (merosin)
o Muscular dystrophy - Fibronectin
o Vascular defects, embryonic lethal (Die in embryo due to inability to form structure of vesicular system) - Fibrinogen
o Bleeding problems, failure of pregnancy (due to failure of placenta)
EMC AND CELL ADHESION
cells attach to ECM via…
adhesion receptors known as integrins
EMC AND CELL ADHESION
at most basic
- At most basic attach cells to EMC (basal lamina)
EMC AND CELL ADHESION
hemidesmosomes and focal adhesions
Hemidesmosomes and focal adhesions are multi-protein complexes that are essential for attachment of epithelial cells to the basal lamina
- Do this through integrins (trans-membrane adhesion receptors)
EMC AND CELL ADHESION
hemidesmosomes
Found in epithelial cells, attaching them firmly to the basal lamina.
Resist shear forces, providing mechanical stability.
Connect keratin filaments inside the cell to laminin in the ECM.
Mutations in components cause skin blistering diseases (e.g., epidermolysis bullosa).
EMC AND CELL ADHESION
focal adhesions
Found in many cell types, especially where cells need to move, spread, or sense their environment.
Connect actin filaments to the ECM via integrins.
Play key roles in:
Cell migration
Signal transduction (via FAK, Src)
Mechanosensing (sensing tension or stiffness in the ECM)
EMC AND CELL ADHESION
hemidesmosomes and focal adhesions
how they function differently
How They Differ Functionally
Hemidesmosomes = anchoring bolts (stability, long-term attachment).
Focal adhesions = traction points (dynamic, adaptable, for movement and signaling).
EMC AND CELL ADHESION
hemidesmosomes
Function
Strong, stable adhesion to basal lamina
EMC AND CELL ADHESION
hemidesmosomes
Main ECM Ligand
Laminin (via integrin α6β4)
EMC AND CELL ADHESION
hemidesmosomes
Key Integrin
α6β4
EMC AND CELL ADHESION
hemidesmosomes
Intracellular Link
Intermediate filaments (e.g., keratin)
EMC AND CELL ADHESION
hemidesmosomes
Cytoskeletal Role
Anchor cells for stability
EMC AND CELL ADHESION
hemidesmosomes
Tissue Location
Epithelia (especially skin, cornea)
EMC AND CELL ADHESION
hemidesmosomes
apperance
Button-like structures at basal surface
EMC AND CELL ADHESION
focal adhesions
Function
Dynamic adhesion involved in signaling and movement
EMC AND CELL ADHESION
focal adhesions
Main ECM Ligand
Fibronectin (via integrin α5β1 or αvβ3)
EMC AND CELL ADHESION
focal adhesions
Key Integrin
α5β1, αvβ3 (others possible)
EMC AND CELL ADHESION
focal adhesions
Intracellular Link
Actin cytoskeleton
EMC AND CELL ADHESION
focal adhesions
Cytoskeletal Role
Coordinate cell adhesion, migration, and signaling
EMC AND CELL ADHESION
focal adhesions
Tissue Location
Most cell types, especially migratory or spreading cells
EMC AND CELL ADHESION
focal adhesions
Appearance
Elongated patches, often at cell edges
EMC AND CELL ADHESION
integrins
Integrin based complexes cluster (into large complexes) to form strong attachment sites for cells
Integrins are heterodimeric transmembrane receptors (α and β subunits) that bind to specific ECM components.
Each integrin has a preferred ligand (or set of ligands), and the binding often involves short peptide motifs like RGD (Arg-Gly-Asp).
EMC AND CELL ADHESION
integrins - the basics
Very abundant on surface of most cells
Alpha subunit
- 18 different alpha subunits
- 3-4 divalent cation binding domains
- Single trans-membrane domain
- Short (<60aa) cytoplasmic tails
Beta subunit
- 8 different beta subunits
- Short (30-50aa) cytoplasmic tails
Form ab heterodimers
- Non-covalently associated
- Low affinity receptors (‘velcro’)
- 24 different ab pairings identified
EMC AND CELL ADHESION
integrins - ligands
fibronectin
- a3b1, a4b1
- Cell adhesion, migration, wound healing; RGD motif
laminin
- a1b1, a6b4
- Basement membrane attachment (e.g. in hemidesmosomes)
collagen
- a1b1, a2b1
- Tissue integrity, structure; important in connective tissue
vitronectin
- avb1, a2b1
-Cell spreading, migration; often seen in serum or wounds
fibrinogen
- avb3, aMb2
- Do not need to memorise
o Just key features:
Alpha and beta subunit
Remember 2 to use as examples
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
- Severity varies
- Important links to embryology
- Remember 1 – 2 example to illustrate importance of integrins
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
alpha 1
- Viable
- No obvious phenotype
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
alpha 3
- Perinatal death
- Kidney tubule defects
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
alpha 5
- Embryonic death
- Cardiovascular defects, neural crest cell death
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
alpha 7
- Viable
- Muscular dystrophy
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
beta 1
- Embryonic lethal
- Failure of gastrulation
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
beta 2
- Viable
- Impaired immune response, persistent infections
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
beta 3
- Viable
- Clotting defects
EMC AND CELL ADHESION
Selected integrin knockout phenotypes
beta 4
- Perinatal death
- Blistering of skin, other epithelial tissues affected
EMC AND CELL ADHESION
Selected integrin knockout phenotypes beta 5
- Viable
- No obvious phenotype
EMC AND CELL ADHESION
SIGNALLING : How integrins regulate cell function via focal adhesions
Two types of signalling involved
- Inside out
- Outside in
EMC AND CELL ADHESION
SIGNALLING : How integrins regulate cell function via focal adhesions
inside out signalling
- Integrins have an active (able to bine to ECM) and inactive (unable to bind to EMC) state and cells can regulate whether an integrin is active or inactive
- This process of regulating integrin activity is know as inside out signalling
o Cell generates signal that tells integrin to switch on or off
ECM binding triggers intracellular pathways:
Signals from inside the cell (e.g. via talin/kindlin) increase integrin affinity for ECM.
Important in immune cells for rapid activation of adhesion.
EMC AND CELL ADHESION
What does activation of an integrin entail
- Involves a structural change
- Cytoplasmic tails linked to cytoskeleton
EMC AND CELL ADHESION
Inside out signalling – an example
- Platelets express alpha IIb beta3 integrin
- On normal circulating platelets, alpha IIb beta3 is inactive
o Does not bind its soluble ligand – fibrinogen - In response to wounding (for example) platelets are activated by thrombin via a G protein-coupled receptor
o Thrombin also cleaves fibrinogen to form fibrin - This results in activation of alpha IIb beta3 such that the platelet can now bind to fibrin and contribute to the clotting response
signals inside cells are changing the conformation of the integrins to switch it on or off
EMC AND CELL ADHESION
outside in signalling
- When active and bound to ECM, integrins can regulate cell function
o E.g. milk production in the mammary gland - This process whereby integrin interaction with ECM regulates cell function is know as outside in signalling
ECM binding triggers intracellular pathways:
Changes in gene expression, cytoskeletal dynamics, survival, migration.
EMC AND CELL ADHESION
outside in signalling - example
- Mammary gland
extracellular signal prolactin
get milk production (cells need on be on laminin)
- Therefore for B-casein production, 2 signals are required:
1) Hormone (prolactin)
2) Laminin interaction with integrin
EMC AND CELL ADHESION
Problem: INTEGRINS HAVE NO ENZYMATIC ACTIVITY
how to they signal
o Cytoplasmic tail sequences provide a clue
- Almost 100% conservation when looking at amino acids of cytoplasmic tails
o (this similarity is not seen else where)
While integrins themselves have no kinase or phosphatase activity, they signal by recruiting and clustering cytoplasmic proteins that do.
Mechanism:
Ligand binding (to ECM) triggers integrin clustering in the membrane.
This clustering leads to formation of focal adhesion complexes.
These complexes recruit and activate signaling molecules.
EMC AND CELL ADHESION
The integrin adhesions network
- These focal adhesions are not just attachment points
o Left with proteins that are in the focal adhesion complex
o Can identify proteins with mass spec - There are known protein kinases
o signalling to the nucleus
o Acting as assembling points for signals to affect cell function - Different pathways different signals
- Recruiting signalling molecules that go onto effect gene regulation
EMC AND CELL ADHESION
collagen - 5 main types
Type I
90% of collagen, found in skin, bones, tendons, and ligaments
EMC AND CELL ADHESION
collagen - 5 main types
Type II
Found in elastic cartilage, provides joint support
EMC AND CELL ADHESION
collagen - 5 main types
Type III
Muscles, arteries and organs
EMC AND CELL ADHESION
collagen - 5 main types
Type IV
Kidneys, ears, eyes
EMC AND CELL ADHESION
collagen - 5 main types
Type 5
Cornea, skin, hair, and placenta
EMC AND CELL ADHESION
collagen - Osteogenesis imperfecta (brittle bone disease)
- Caused by a mutation in the gene for collagen type I
o COL1A1 or COL1A2
o 90% of collagen is type 1
o Mutations have knock on (a big) effects - As with all the inherited collagen diseases, inherited as a dominant trait
- Even though one collagen allele is normal, the assembly of the normal gene product with the mutant product result in defective collagen fibres
o Structure gives strength to tissue
o The fact that one allele is mutation end up with defective collagen fibres
o End up with weakened collagen (usually in long bones)
EMC AND CELL ADHESION
collagen - aplort syndrome
symptoms
o Kidney function is impaired (haematuria, proteinuria, hypertension); hearing is impaired (in adults); myopia (short-sightedness)
EMC AND CELL ADHESION
collagen - aplort syndrome
mutations
- Mutations in collagen IV biosynthesis genes
o COL4A3, COL4A4, COL4A5 - Can be X-linked (COL4A5, 85% of cases) or autosomal recessive (COL4A3, COL4A4)
- Defective collagen IV assembly leads to impaired filtration of urine and can eventually lead to kidney failure
EMC AND CELL ADHESION
collagen - aplort syndrome
the basal lamina
The principle components are:
Collagen type IV
* Forms grid with all other things assemble around
* And directly interacts
* If ordered collagen is disrupted you get a basal lamina that doesn’t quite have the integrity it should have
Nidogen
Laminin
Perlecan (a proteoglycan)
The basal lamina acts as a molecular filter preventing passage of macromolecules
- E.g. proteins from the blood to urine
EMC AND CELL ADHESION
collagen - aplort syndrome
the kidney
- This is the unit that acts as a filtration
- Inserted between the blood vessels and the collecting ducts is a basal lamina
- Prevents macromolecules such as proteins from passing into the urine
- If basal lamina has holes in it (because collagen not assembled properly)
these macromolecules can get through
EMC AND CELL ADHESION
Epidermolysis Bullosa (EB)
- EB is the generic term is describe a number of different skin diseases that share certain similarities
- Inherited
- Involve increased skin fragility
o Prone to blistering and damage - Blistering of skin and mucous-secreting epithelial cells including those that line the gut
- Three main types:
o Epidermolysis bullosa simplex (70%)
o Junctional EB (10%)
o Dystrophic EB (20%)
EMC AND CELL ADHESION
Epidermolysis Bullosa (EB)
The human epidermis is a stratified epithelial tissue that forms…
the outermost protective layer of the skin and is comprised predominantly of cells known as keratinocytes
EMC AND CELL ADHESION
Epidermolysis Bullosa (EB)
through skin; looking at epidermis
- As cells are lost have to replace them
- Cells are lost from the surface of your skin all the time
- Division and differentiation (series of biochemical and morphological changes) at the bottom (the basal layer of the epidermis)
- Move up to conifer cells on the surface
o These are the protective cells
EMC AND CELL ADHESION
Epidermolysis Bullosa (EB)
how does a blister form
- Bilsters are formed by friction
- Causes sheer forces between the epidermis and the underlying dermis
- And connecting these is the basement membrane
- Can rupture the connection between epidermis (basal layer) and basement membrane
o They detach
o This forms a blister (fills with fluid)
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
- Inherited condition
- Characterised by structural and mechanical fragility of skin and mucosal surfaces
- Two main types (features overlap, and can be caused by mutations in same genes)
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
o Generalised intermediate JEB (non-herlitz JEB)
Lifelong pain and disability
Increased incidence of skin cancer
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
o Generalised severe JEB (herlitz JEB)
Frequently fatal during first 1-2 years
Due to malnutrition and anaemia as a consequence of severe blistering in the pharynx and oesophagus
* Internal mucosal epithelial linings affects
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
genetics
- Can result from mutations in the following genes:
o LAMB3 – encodes laminin subunit beta3
o LAMA3 – encodes laminin subunit alpha3
o LAMC2 – encodes laminin subunit gamma2
Involved the extracellular matrix molecule laminin (laminin 332 - used to be known as 5)
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
laminin
Laminin is an important component of the basement membrane to which keratinocytes attached via integrins
EMC AND CELL ADHESION
Junctional Epidermolysis Bullosa (JEB)
mutations affect binding to integrins
Key binding subunit
Needs to be assembled properly
integrins involved in laminin binding
Alpha 6 beta 4
* Found in hemidesmosomes
Alpha 3 beta 1
* found on…?focal? adhesions
Gene therapy: has been used to successfully treat some individuals with EB
- ‘butterfly child’
EXTRACELLULAR MATRIX PROTEIN – ELASTIC FIBRES
- To function correctly many tissues (e.g. skin, blood vessels) need to be both strong and elastic
o Need to avoid over stretching these structures - Strength comes from collagen
- Elastic fibres give tissues their elasticity
EXTRACELLULAR MATRIX PROTEIN – ELASTIC FIBRES
elastin
- A very hydrophobic protein (many prolines and glycines)
- Cross-linking of individual molecules of elastin results in a network of elastic fibres (amorphous elastin) that are 5 times more extendable than a rubber band of equivalent cross section
EXTRACELLULAR MATRIX PROTEIN – ELASTIC FIBRES
elastic fibres - importance of fibrillin
- Over-extension is prevented as Fibrillin fibres microfibrils form a scaffold surrounding the amorphous elastin
- Need to prevent overstretching
- if stretch too far will snap
- Prevented in arteries using another extracellular matrix protein
o Fibrillin - Have a core of elastin fibres
- Surrounding them have rope like structures of fibrillin
o Doesn’t stretch
EXTRACELLULAR MATRIX PROTEIN – ELASTIC FIBRES
Marfan’s syndrome: clinical features
o Tall stature with long limbs and fingers (arachnodactyly)
o Lens defects (not seated properly in eye, tends to get dislocated)
o Aortic aneurysm (prior to genetic screening, those undiagnosed suffer)
EXTRACELLULAR MATRIX PROTEIN – ELASTIC FIBRES
Marfan’s syndrome: genetics
o Autosomal dominant
o Mutation in FBN1 gene
o FBN1 encodes Fibrillin not assembled correctly, so over time stretches far
definition of ECM
chatgpt
The ECM is a dynamic, complex network of secreted proteins and polysaccharides that not only provides structural support to tissues, but also actively influences cell behavior, including migration, differentiation, proliferation, and survival.
Main Functions of ECM:
chatgpt
Structural framework for tissue integrity
Barrier/filter (e.g., basal lamina in kidney)
Signal reservoir (binds growth factors and presents them to cells)
Cell migration guidance (during development and wound healing)
Regulation of gene expression via integrin-mediated signaling
Mechanical properties (e.g., elasticity via elastin/fibrillin)
Hemidesmosomes
Associated diseases
chatgpt
Epidermolysis bullosa
Focal Adhesions
Associated diseases
Cancer metastasis, wound healing defects
Inside-Out signaling
chatgpt
Intracellular signals switch integrins from inactive to active
e.g. Platelet αIIbβ3 activation by thrombin
outside in signalling
chatgpt
ECM binding triggers intracellular signaling via adaptors
e.g. Laminin–integrin → milk production
signalling is modulated by…
Signaling is modulated by ECM stiffness, ligand density, and receptor clustering.
SIGNALLING: How integrins regulate cell function
When integrins bind ECM components like laminin or fibronectin, they cluster and recruit signaling complexes (e.g., FAK, Src, paxillin).
SIGNALLING: How integrins regulate cell function
what pathways do integrin binding trigger
MAPK/ERK → proliferation
PI3K/Akt → survival
Rho GTPases → cytoskeletal changes/migration
YAP/TAZ → mechanosensing and gene regulation
SIGNALLING: How integrins regulate cell function
what does this enable integrins to do
Control cell fate decisions (e.g., differentiation vs. proliferation)
Regulate gene expression
Coordinate cell movement and shape
Respond to mechanical forces in the ECM
