The extracellular environment Flashcards
What is an extracellular matrix (ECM)?
- Secreted molecules that constitute the cell microenvironment
- E.g. Glycoproteins, collagens, glycosaminoglycans and proteoglycans.
What are some of the functions of the ECM?
- Provides the bulk, shape and strength of tissues in vivo e.g. bone
- The substrate for cell migration
- Provides the spacial context for signalling events including growth factor receptors and adhesion molecules
- Can determine cell behaviour e.g. polarity, migration and differentiation, by communicating with intracellular cytoskeleton and transmitting growth factor signals
Why must cells be supported structurally by the ECM?
- Because cells can’t function properly without structural support e.g. anoikis
- Anoikis - A programmed cell death that occurs when cells anchorage-dependent cells, e.g. epithelial cells, detach from the ECM
Explain how the ECM acts to provide support cells
- Forms the basement membranes in the skin to provide anchoring support, tensile strength and flexibility
- ECM proteins, e.g. integrins, can serve as ligands for focal adhesion
- ECM proteins can also bind to other ECM components which contributes to the structure of the ECM
What produces the ECM?
- The ECM is produced by stromal cells within the matrix itself e.g. fibroblasts, osteoblasts and epithelial cells.
Which properties of the ECM determine cell behaviour and morphology?
- Type of ECM - e.g. rigid or soft
- Physical properties - e.g. what proteins make up that ECM
Name the 5 major ECM proteins
- Collagens
- Laminins
- Fibronectin
- Vitronectin
- Elastin
Describe some characteristics of collagens
- Primary proteins present in the ECM and basememt membrane
- At least 29 members
- Types I, III, IV and XI are the major members
- Provide structural support and binding partners for other ECM proteins
- Insoluble glycoproteins
- Proline and hydroproline make up 1/6 of protein sequence and provide stiffness to polypeptide chain
Describe some charcteristics of Laminins
- Mainly found in the basement membrane
- Provide tensile strength to tissue
- 3 subunits:
- Alpha (5 genes)
- Beta (4 genes)
- Gamma (3 genes)
- These 3 subunits form characteristic cross pattern
Describe some characteristics of fibronectin
- Exists as a dimer
- Can bind to collagens and heparan sulphate proteoglycans.
Describe some characteristics of vitronectin
- Binds to and regulates the components of the urokinase plasminogen activator (uPA) complex
What is the urokinase plasminogen activator (uPA) and what is its function?
- uPA is a serine protease involved in the degradation of the ECM
- It cleaves plasminogen into plasmin which then cleaves pro-MMPs into MMPs (matrix metalloproteinases)
- Also partcipates in cell adhesion through its association with integrin
Describe some characteristics of Elastin
- Major structural protein in the ECM
- Individual tropoelastin subunits are crosslinked to give the mature elastin fiber
- Responsible for flexibilty in many tissue along with fibrillin
What are proteoglycans?
- A class of protein where the core protein is decorated with covalently linked glycosaminoglycan chains
What are the 3 types of proteoglycans?
- Heparan sulphate proteoglycans (HSPGs)
- Chondroitin sulphate proteoglycans (CSPGs)
- Hyaluronic acid
Describe some charcateristics of heparan sulphate proteoglycans (HSPGs)
- Consist of a core protein covalently linked attached to heparan sulphate (HS) chains
- HS is a linear polysaccharide with a repeating dissacharide unit backbone
- Each disaccharide unit undergoes specific mofification patterns to regulate its function e.g. addition of sulphate groups
Give some examples of heparan sulphate proteoglycans
- Perlecan
- Agrin
- Syndecan
- Glypicans
What are the functions of heparan sulphate proteoglycans?
- Bind to the other ECM structural proteins (e.g. laminin, fibronectin, collagen) via heparan sulphate side chains.
- Control the diffusion of the growth factors to establish morphogen gradients in development
- Act as a respository for growth factors. e.g. VEGFs, which they then release at an appropriate time (e.g. wound healing).
Describe some characteristics of Chondroitin sulphate proteoglycans (CSPGs)
- Long, unbranched chondroitin sulphate chains (disaccharide unit repeats) attached to a protein core
- Examples of CSPGs are: Aggrecan, versican, decorin, biglycan
- Can be modified (by addidition of Chondroitin sulphate) so you have four different disaccharide unit subtypes bearing one or two negatively charged sulphate groups in different positions.
What are the functions of Chondroitin sulphate proteoglycans
- Bind and crosslink with other ECM components
- Regulate the growth factors (e.g. TGF) just like HSPGs
Describe some characteristics of hyaluronic acid (including its function)
- A glycosaminoglycan consisting of alternating glucuronic acid (red diamond) and N-acetyl glucosamine (blue square) sugars.
- Responsible for the gel like tissues such as cartilage
As well as proteins and proteoglycans the ECM also contains extracellular proteases. Why does the ECM contain extracellular proteases?
- Because in order to allow for the free movement of cells or deposition of new matrix, ECM must be degraded by extracellular proteases
What are the different types of extracellular proteases?
- Matrix metalloproteinase (MMP) families
- Metalloproteinase with thrombospondin motif (ADAMTS) families
- Heparanase
- Chondroitinase
What is the FGFR (fibroblast growth factor receptor) and what function does HSPG have within the recepetor?
- Member of receptor tyrosine kinase family (FGFR 1-4)
- Alternative splicing of Ig II & III domains creates ligand binding specificity (isoforms a-c)
- Heparan sulphate (HS) is essential for the stabilisation and functional activation of the receptor
ECM proteins such as fibronectin have multiple domain stuctures, what is the function of these domain stuctures?
- Fibronectin exists as a dimer and can bind to other ECM proteins via its various binding sites
- These binding sites are formed by the multiple domain stuctures that Fibronectin and other ECM proteins have e.g. FN3
- Fibronectin is encoded by a single gene but different domains are formed through alternative splicing
- Binding of multiple ECM proteins to fibronectin allows for multi-domain interactions between the different proteins
What is the use of the multi-domain interactions of fibronectin?
- FN organizes and integrates all the various signals from these interactions at three levels.
- recruits growth factors to the ECM
- physically localizes the receptors on the cell surface
- organises the different receptors and domains into a submicron patch in the membrane, so the receptors will be brought into close proximity such that their signals provide integrated information to the cell, generating “chords” and “melodies” in contrast with the “single notes” generated by each receptor.
What are focal adhesion sites?
- Focal adhesion sites are structures that connect the ECM to the cytoskeleton of a cell (specifically actin)
- They’re formed right above the cytoskeletal bundle that connects the intracellular cytoskeleton
What is the specific function of focal adhesion sites
- Anchor bundles of actin stress fibers (F-actin) through many proteins, including αvβ3 integrins and structural proteins like alpha-actinin, vinculin and talin
What is the importance of the Interconnection of the actin machinery and focal adhesions?
- This interaction allows for the cooperation of several components at the focal adhesion sites and this forms the cell’s contractibility and mechano-responsive network, including:
- Actin polymerization and myosin II-dependent machinery
- Specific linking proteins (e.g. talin and vinculin)
- The cell surface adhesion receptors (e.g. integrins)
- The co-receptors (e.g. heparan sulphate proteoglycans, such as syndecans).
How are integrins activated and what happens as a result of this activation?
- Activated via dimerisation of α–β subunits and ligation of those subunits to the ECM which initiates focal adhesion formation.
Do integrins have alternatively spliced forms?
- Yes and each spliced form has specific ligands it can bind to
Explain how trafficking of focal adhesion molecules regulates directional cell movement
- Integrin endocytosis from the rear to the front of the cell regulates directional cell movement.
- Integrin molecules, bound to the ECM, become endocytosed into early endosomes (EE), transported to the recycling endosome (RE) and inserted into the membrane at the cell front.
- At the front of the cell, integrin molecules become internalized, which constantly removes them from the plasma membrane so that they cannot diffuse laterally along the membrane (Integrin endocytosis counteracts lateral diffusion).
What is the epithelial to mesenchymal transition (EMT)?
- The transition from a cuboidal, attached phenotype (epithelial) to an elongated, mobile conformation (mesenchymal).
What growth factor signalling pathways occur at focal adhesion sites and what role does the ECM have in these signalling pathways?
- The central signalling pathway is the Src/FAK complex, which activates ERK, Akt, RhoA, Rac1 and JNK to regulate cell survival, proliferation, and differentiation.
- ECM regulates the growth factor downstream signalling in cooperation with integrins and growth factor receptors.
Apart from mediating cell survival and growth as part of integrin signalling what else can the Src-FAK complex do?
- Src–FAK complex can also integrate their signals with the signals produced by growth factors signalling pathways.
What are the effects of the cooperation between integrin signallign and growth factor signalling?
- Integrin ligation may lead directly to the increased secretion of growth factors which can then bind to their receptors in an autocrine or paracrine manner to further induce signalling.
- Signalling induced by either integrin ligation or growth factor binding may activate common downstream pathways, resulting in enhanced signalling overall, compared with the activation of either receptor alone.
- Chemokine and growth factor signalling may regulate integrin function by directly controlling integrin expression levels.
What processes do integrins expressed in tumour cells facilitate and how do they do this?
- Facilitate tumour progression and metastasis by increasing tumour cell migration, invasion, proliferation and survival.
Give an example of an ECM protein whose expression is affected by cancer
- Endostatin in human breast cancer
-
In normal breast tissue
- The endostatin (red) is located in the basal membrane underlining the ductal epithelium, as well as the blood vessels (CD31, green).
- Nucleus is shown in blue (DAPI stain).
-
In breast cancer
- The endostatin expression is lost from the epithelium and a diffused signal is now seen in the tumor stroma.
- NOTE: Endostatin is an inhibitor of angiogenesis
Give some examples of therapeutic agents that target the ECM. DON’T memorise table
