Extracellular Matrix 2
- 1 Extracellular Matrix 2
- 2 Introduction
- 3 Objectives
- 4 Proteoglycans
- 5 Fibronectin
- 6 Laminin
- 7 Basement Membrane
- 8 Basal Lamina Experiment
- 9 ECM Reorganisation
- 10 History
- 11 ECM Scaffold - Tissue Engineering
- 12 References
- 12.1 Recent Reviews
- 12.2 Textbooks
- 12.3 Books
- 12.4 PubMed
- 13 Images
- 14 2016 Course Content
Extracellular Matrix 2
This second lecture on ECM will cover the glycoprotein components of ECM and specialized epithelial ECM. Finally I will discuss some key experiments exploring the role and function of the ECM of epitheilia (basement membrane) and connective tissues. With the epithelial ECM the term "basement membrane" is used with light microscopy and "basal lamina" is used with electron microscopy.
- due to the 2016 Monday public holiday - I will be continuing into ECM2 and also covering this content in this week's practical class.
- 2015 ECHO Recording ECM 2
- Last year's student projects were designed around ECM - 2015 Extracellular Matrix: Group 1 - Small Leucine-Rich Proteoglycans | Group 2 - Integrins | Group 3 - Elastic Fibres | Group 4 - Fibronectin | Group 5 - Laminin | Group 6 - Collagen | Group 7 - Basement Membrane
MH - note that content listed below will not match exactly current lecture structure but has been selected as having similar content
- Understanding of the localisation and origin of extracellular matrix
- Understanding of the 3 major components
- fibers, proteoglycans (matrix), adhesive glycoproteins
- Broad understanding of structure and function
- basement membrane
- Broad understanding of some key extracellular matrix experiments
- consist of protein (~5%) and polysaccharide chain (~95%)
- form a gel to embed the fibril network
- Golgi apparatus - GAG disaccharides are added to protein cores to form proteoglycans
- 10% by weight but fill most of space
- unbranched polysaccharide chains
- disaccharide subunits
- amino sugar
- Hyaluronan (or hyaluronic acid) main glycosaminoglycan in connective tissue
- high molecular weight (~ MW 1,000,000 )
- length of about 2.5 µm hyaluronan
- "backbone" for the assembly of other glycosaminoglycans
- Hyaluronan is also a major component of the synovial fluid, which fills joint cavities, and the vitreous body of the eye.
- Other 4 major glycosaminoglycans
- chondroitin sulphate, dermatan sulphate, keratan sulphate and heparan sulphate (UK sulphate, US sulfate)
- attach through core and link proteins to hyaluronic acid backbone
- differs from other GAG synthesis
- synthesized at plasma membranes
- nascent chains directly extruded into ECM
Proteoglycan - Disease
Mucopolysaccharidosis type I (MPS I) - Hurler disease
- deficiency of alpha-L iduronidase => lysosomal storage disease, associated with an altered elastic matrix
- excess heparan sulphate and dermatan sulphate
- altered types and kinds of proteoglycans formed by cells
- normal cells -> malignant
- Cartilage breakdown (cartilage erosion)
- chondrocytes elicit a catabolic response which exceeds anabolism of new matrix molecules
- Degrade proteoglycan (aggrecan)
- Also a mouse model generates antibodies to proteogycan
- dimer connected at C-terminus
- Mr 550 kDa
- nearly identical subunits composed of types I (F1), II (F2), and III (F3) fibronectin modules
- S-S linkages
- rigid and flexible domains
- fibronectin fibrils have elastic properties and can stretch fibrils up to four-fold their relaxed length.
- fibrillogenesis - transformation from the compact (soluble) form to the extended fibrillar (insoluble) form of fibronectin, requires application of mechanical forces generated by cells.
- cell binding segment RGDS
- binds integrin receptor in membrane
- then mechanically couples to the actin cytoskeleton
- domains bind
- heparin sulphate
- hyaluronic acid
- soluble protein in blood plasma (200–250 kDa monomer)
- blood clotting process, link to fibrin
- insoluble protein in extracellular matrix (ECM)
- ECM fibronectin differs from plasma fibronectin by the presence of additional polypeptide segments and in altering morphology of transformed cells and hemagglutination.
- blocking fibronectin with antibody
- prevents neural crest migration
- extension of axons and dendrites
Fetal fibronectin (fFN)
- produced by fetal cells and found acting as an adhesive at the interface of the chorion and the decidua ( fetal membrane and uterine lining).
- has been used as a diagnostic for preterm birth.
- cell adhesion
- migration pathways
- stimulates growth of axons
- development and regeneration
- basal laminae
- most abundant linking glycoprotein
Integrin- Structure Integrin Function cell membrane receptor for ECM linkers binds RGDS motif 2 subunits alpha (α) and beta (β) transmembrane linked to cell cytoskeleton actin microfilaments via talin and vinculin focal contacts For Review see Integrin signaling revisited. Schwartz MA.Trends Cell Biol 2001 Dec;11(12):466-70
Integrin and Laminin - Several integrin heterodimers act as laminin receptors on a variety of cell types alpha 1 beta 1 alpha 2 beta 1 alpha 3 beta 1 alpha 6 beta 1 alpha 7 beta 1 alpha 6 beta 4 Microsc Res Tech 2000 Nov 1;51(3):280-301
Integrin and Laminin
- Roles of laminin-binding integrins in adhesion-mediated events in vertebrates
- embryonic development, cell migration and tumor cell invasiveness, cell proliferation, differentiation and basement membrane assembly
- essential role for receptors in maintaining cell polarity and tissue architecture
The epithelial ECM the term "basement membrane" is used with light microscopic observation and "basal lamina" is used with electron microscopy.
The basement membrane is composed of two sublayers.
- (about 40–120 nm thick) consists of fine protein filaments embedded in an amorphous matrix.
- Membrane proteins of the epithelial cells are anchored in the basal lamina, which is also produced by the epithelial cells.
- major component of the basal lamina are two glycoproteins - laminin and (usually type IV) collagen
- consists of reticular fibres embedded in ground substance.
- fibres of the reticular lamina connect the basal lamina with the underlying conective tissue.
- components of the reticular lamina are synthesised by cells of the connective tissue underlying the epithelium.
Basal Lamina Experiment
Figure 19-60. Regeneration experiments demonstrating the special character of the junctional basal lamina at a neuromuscular junction this link currently not functional. temp link
- Basal lamina directs acetylcholinesterase (AChE) accumulation at synaptic sites in regenerating muscle
- skeletal muscle damaged such that basal lamina sheaths of the muscle fibers spared
- new myofibers develop within sheaths and neuromuscular junctions form at original synaptic sites
- regenerated neuromuscular junctions have junctional folds and accumulations of acetylcholine receptors and AChE
|Neuromuscular Junction Expt|
- Activate members of Rho-family of small GTPases
- Conversely, Rho- and Ras-family proteins can influence the ability of integrins to bind their ligands
- control of cell motility, and therefore of invasive and metastatic behavior
- Integrin binding ECM has effects on cell survival, particularly for cells of epithelial origin
- specific integrins have selective effects on efficiency of signal transduction in cell survival pathways
Text modified from: New aspects of integrin signaling in cancer. Semin Cancer Biol 2000 Dec;10(6):407-14
Reorganisation can occur through proteolytic degradation changes to ECM proteins (collagen, laminin, and fibronectin). Their activity can be regulated locally by inhibitors.
The proteases form 2 main classes:
Matrix Metalloproteases (MMPs)
- dependent upon bound Ca2+ or Zn2+ for activity.
- family of enzymes
- MMP-2 (Gelatinase A, 72 kDa type IV collagenase) is the most widely distributed
- collagenases can specifically cleave proteins at a small number of sites.
- inhibited by tissue inhibitors of metalloproteases (TIMPs).
- MMP-2 appears to be associated with early breast carcinoma and cervical neoplasia
- have a highly reactive serine in their active site.
- inhibited by serpins.
- role in metastasis
Below are some example historical research finding related to cell junctions from the JCB Archive.
1978 Basal lamina instructs innervation Joshua Sanes and Jack McMahan show that regenerating nerve axons take their cues for new synapse formation from the extracellular matrix (ECM) of muscle cells and not from the muscle cells themselves.
ECM Scaffold - Tissue Engineering
- “Decellularized tissues and organs have been successfully used in a variety of tissue engineering/regenerative medicine applications, …Each of these treatments affect the biochemical composition, tissue ultrastructure, and mechanical behavior of the remaining extracellular matrix (ECM) scaffold, which in turn, affect the host response to the material.”
Reference: Decellularization of tissues and organs. Biomaterials. 2006 Jul;27(19):3675-83. Epub 2006 Mar 7.
<pubmed>26900407</pubmed> <pubmed>25487405</pubmed> <pubmed>26635793</pubmed> <pubmed></pubmed>
Essential Cell Biology
- Essential Cell Biology Chapter 19 p594-604
Molecular Biology of the Cell
Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter New York and London: Garland Science; c2002 --Mark Hill (talk) 08:58, 1 May 2014 (EST) These library links currently not functional.
- Molecular Biology of the Cell 4th ed. - V. Cells in Their Social Context Chapter 19. Cell Junctions, Cell Adhesion, and the Extracellular Matrix
- The Extracellular Matrix of Animals
- Figure 19-53. The structure of a fibronectin dimer
- Figure 19-54. Coalignment of extracellular fibronectin fibrils and intracellular actin filament bundles
- Figure 19-57. The structure of laminin
- Figure 19-58. A model of the molecular structure of a basal lamina
- Figure 19-60. Regeneration experiments demonstrating the special character of the junctional basal lamina at a neuromuscular junction
Molecular Cell Biology
Lodish, Harvey; Berk, Arnold; Zipursky, S. Lawrence; Matsudaira, Paul; Baltimore, David; Darnell, James E. New York: W. H. Freeman & Co.; c1999
- Molecular Cell Biology - Chapter 22. Integrating Cells into Tissues
- Cell-Matrix Adhesion
- Figure 22-22. Structure of fibronectin chains
The Cell- A Molecular Approach
Cooper, Geoffrey M. Sunderland (MA): Sinauer Associates, Inc.; c2000
- The Cell- A Molecular Approach
- The Cell - A Molecular Approach - III. Cell Structure and Function Chapter 12. The Cell Surface
- The Extracellular Matrix
Essentials of Glycobiology, 2nd ed.
Varki, A.; Cummings, R.D.; Esko, J.D.; Freeze, H.H.; Stanley, P.; Bertozzi, C.R.; Hart, G.W.; Etzler, M.E., editors Plainview (NY): Cold Spring Harbor Laboratory Press; 2008
Search Online Textbooks
- "extracellular matrix" Molecular Biology of the Cell | Molecular Cell Biology | The Cell- A molecular Approach | Bookshelf
- "fibronectin" Molecular Biology of the Cell | Molecular Cell Biology | The Cell- A molecular Approach | Bookshelf
- "laminin" Molecular Biology of the Cell | Molecular Cell Biology | The Cell- A molecular Approach | Bookshelf
- "basement membrane" Molecular Biology of the Cell | Molecular Cell Biology | The Cell- A molecular Approach | Bookshelf
- "basal lamina" Molecular Biology of the Cell | Molecular Cell Biology | The Cell- A molecular Approach | Bookshelf
- PubMed is a service of the U.S. National Library of Medicine that includes over 18 million citations from MEDLINE and other life science journals for biomedical articles back to 1948. PubMed includes links to full text articles and other related resources. PubMed
- PubMed Central (PMC) is a free digital archive of biomedical and life sciences journal literature at the U.S. National Institutes of Health (NIH) in the National Library of Medicine (NLM) allowing all users free access to the material in PubMed Central. PMC
- Online Mendelian Inheritance in Man (OMIM) is a comprehensive compendium of human genes and genetic phenotypes. The full-text, referenced overviews in OMIM contain information on all known mendelian disorders and over 12,000 genes. OMIM
- Entrez is the integrated, text-based search and retrieval system used at NCBI for the major databases, including PubMed, Nucleotide and Protein Sequences, Protein Structures, Complete Genomes, Taxonomy, and others Entrez
- "extracellular matrix" Entrez all databases
- "fibronectin" Entrez all databases
- "laminin" Entrez all databases
- "basal lamina" Entrez all databases
- "basement membrane" Entrez all databases
- "matrix metalloprotease" Entrez all databases
- "serine proteases" Entrez all databases
- note the spelling differences when carrying out other related ECM searches: UK sulphate, US sulfate ; UK fibre, US fiber
- Hay ED. The extracellular matrix in development and regeneration. An interview with Elizabeth D. Hay. Int J Dev Biol. 2004;48(8-9):687-94. No abstract available. PMID: 15558460
- Hay ED. Extracellular matrix. J Cell Biol. 1981 Dec;91(3 Pt 2):205s-223s. Review. No abstract available. PMID: 6172429
- Sasaki T, Fässler R, Hohenester E. Laminin: the crux of basement membrane assembly. J Cell Biol. 2004 Mar 29;164(7):959-63. Epub 2004 Mar 22. Review.
2016 Course Content
Lectures: Cell Biology Introduction | Cells Eukaryotes and Prokaryotes | Cell Membranes and Compartments | Cell Nucleus | Cell Export - Exocytosis | Cell Import - Endocytosis | Cytoskeleton Introduction | Cytoskeleton - Microfilaments | Cytoskeleton - Microtubules | Cytoskeleton - Intermediate Filaments | Cell Mitochondria | Cell Junctions | Extracellular Matrix 1 | Extracellular Matrix 2 | Cell Cycle | Cell Division | Cell Death 1 | Cell Death 2 | Signal 1 | Signal 2 | Stem Cells 1 | Stem Cells 2 | Development | 2016 Revision
Laboratories: Introduction to Lab | Microscopy Methods | Preparation/Fixation | Cell Knockout Methods | Cytoskeleton Exercise | Immunochemistry | Project Work | Confocal Microscopy | Tissue Culture | Stem Cells Lab | Microarray Visit
Dr Mark Hill 2015, UNSW Cell Biology - UNSW CRICOS Provider Code No. 00098G