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1. Aggrecan is a proteoglycan found in cartilage that helps with mechanical support, forms
aggregates w/ hyaluronic acid, and binds TGF- to inhibit ECM synthesis. Gives
cartilage its lubricating characteristic.
2. Cartilage: very rich in GAGs. Cartilage proteoglycan Aggrecan with hyaluronic acid
forms a complex that secures lots of GAGs in the tissue, serves as shock absorber for
compression.
3. Cartilage degeneration is a dominant phenotype of osteoarthritis (OA). GAGs get
broken down leading to less joint lubrication, causes bone-on-bone pressure later on.
Transmembrane Proteins that Mediate Cell Adhesion
1. CAMs glue cells to one another
a. Homophilic Interacting CAMs:
i. Cadherins (E-Cadherins): present in desmosomes and mediate adhesion
between cells during development, allowing tissue formation.
ii. Ig-Superfamily NCAMs: important for neuronal development.
b. Heterophilic Interacting CAMs:
i. Integrins: bind to multiadhesion matrix protein (Fibronectin).
1.
Integrins are obligate heterodimers of and subunits. There are ~ 19 subunits and
8 subunits, and the nature of these determine the integrins binding specificity to
the fibronectin RGD motif, collagen, laminin, or leukocytes.
2.
Integrins also function by interacting with elements the ECM, the cytoplasmic domain
can mediate various intracellular signals, which can define cellular shape, motility, and
even regulate the cell cycle.
3.
Integrins communicate biochemical and mechanical signals in a bidirectional manner
across the PM.
4.
Integrins can mediate indirect interactions with the cytoplasmic cytoskeleton through
intermediary proteins including Talin and Vinculin.
5.
Integrins can also signal within cell through the Paxillin-Focal Adhesion Kinase
(FAK) Complex, which can mediate signals to the Ras-MAP Kinase pathway. Ex:
polymerization of Actin Stress Fibers.
6.
In addition to the GF-receptor pathway, many cell types also require interactions with a
defined ECM to proliferate. Without Integrin signaling the active GF signaling molecules
are short lived. The integrins can also activate the ERK1/2, Phosphatidyl Inositol 3
Kinase (PI3K) pathway, and Rac/Rho/Cdc42 G-proteins. Combined these 2 pathways
permit reorganization of cytoskeleton, contribute to cell proliferation, survival,
migration, and gene transcription.
ii. Selectins (P-Selectin): Carbohydrate-binding proteins that bind to
glycoproteins from other cells.
1. Selectins are important for WBC extravasation. Resting state WBC
express specific glycoproteins that are selectin ligands. Platelet
Activating Factor (PAF) is an inflammatory factor expressed by
platelets, injured tissues, endothelial cells, and some WBCs, and PAF
activates the capillary endothelial cells to cause exocytosis of Pselectin onto the cell surface. The WBC selectin ligand then binds to
the endothelial P-selectin, and causes the RBC to slow down in the
blood and start rolling along endothelial surface. This rolling brings
the WBC close enough to allow PAF to bind to a PAF Receptor on the
WBC, which activates an integrin on the surface of the WBC, causing
firm adhesion of the cells due to binding of both integrins and CAMs
(ICAM). Extravasation occurs between two endothelial cells. Once in
the tissue, the WBC can migration along a chemotactic concentration
gradient of PAD or other chemotactic factor to site of injury.
a. Disruption of this process can decrease inflammation and
serves as a new basis for treatment of MS. In untreated MS
2. G-actin (globular) monomer and F-actin (filamentous). ATP and Mg2+ are REQUIRED for
polymerization of actin. G-actin has binding site for ATP, this cleft is exposed at the
minus (-) end. The end of each monomer binds to the + end of the next monomer,
which gives the filament polarity.
3. Actin-nucleating proteins and actin-binding proteins control the organization of F-actin
into higher order structures.
a. Actin Nucleating and Binding proteins
i. Cofilin: binds to F-actin who subunits contain ADP, and causes the actin
filament to break into shorter pieces, which creates more ends and
enhances disassembly of F-actin.
ii. Profilin: Enhances exchange of ADP for ATP on G-actin.
iii. Thymosin-4: binds to ATP-G-actin and sequesters it from the
polymerization process. It helps to maintain a steady state level of ATP-Gactin for polymerization at the + end of F-actin.
iv. CapZ: binds at the + end of F-actin and prevents further addition of ATP-Gactin monomers. Activity of CapZ is tightly controlled by signal transduction
molecules in the cell and by other regulatory proteins at the + end.
v. Tropomodulin: binds at the end and stabilizes F-actin.
vi. Formin: stimulates the assembly of long unbrached F-actin that is found
in stress fibers and contractile rings in the cell.
vii. Arp2/3 complex: Stimulates formation of branched F-actin filaments,
which are found in the leading edge of migrating cells.
4. Treadmilling: G-actin in ATP form is added to + end (Note: G-actin can polymerize ONLY
in the ATP state). In theory the subunits could bind at either the plus or minus end, but
bind mainly at the + end because it has a lower critical concentration for ATP-G-actin
monomers, thus F-actin grows from the + end. When the ATP-G-actin binds to the +
end, the ATP is hydrolyzed to ADP & Pi and the release of Pi causes a conformation shift
of ADP-G-actin such that it promotes dissociation of the monomer.
5. Cell Migration
a. Steps
i. Extension of lamellipodium (little feet) at leading edge, extension of Factin is nucleated by the Arp2/3 complex, and is controlled by Cdc42 and
Rac (two signaling GTP-binding proteins activated by the GF signaling
cascade).
ii. Adhesion at leading edge
iii. Translocation: shifts cytoplasm forward into leading edge.
iv. De-adhesion: of rear end.
b. GFs often serve as initial chemotactic molecule.
c. At the trailing edge Rho (another GTP-binding protein) signals the activation of
Formin leading to stress fiber formation. Rho also activates Rho Kinase, which
activates Myosin II causing the back of the cell to contract. Rho activation blocks
Rac activation thus establishing polarity in the cell, coordinating events at both
ends.
6. Cross-linking proteins also organize F-actin into networks
a. Fibrin: cross-links parallel F-actin filaments in the intestinal microvilli, and helps to
stabilize the F-actin network to provide PM structure.
b. -actinin organizes the F-actin of the stress fibers during migration. Also
important for muscle contraction.
c. Spectrin: cross-links and organizes F-actin into meshwork just below PM to
support and strengthen the PM. Spectrin and Dystrophin are both in the Spectrin
superfamily of proteins.
7. Integrins and Focal Adhesion:
a. Integrins crosslink the cytoskeletal proteins to the ECM. Integrins FIX cells into
their tissues, establishing focal adhesions.
b. Integrins also help generate intracellular signals that lead to the formation of the
focal adhesions.
8. Hereditary Spherocytosis: RBCs become small and fragile due to defect in spectrin,
protein 4.1, or ankyrin. Patients have anemia and splenomegaly. Gallstones composed
of bilirubin (from RBC breakdown) are also common.
a. Cell fragility is similar to DMD in muscle cells. Dystrophin and Spectrin from same
superfamily.
Intermediate Filaments:
1. Classes
a. I: Acidic keratins. Epithelial cells. Tissue strength and integrity.
b. II: Basic keratins. Epithelial cells. Tissue strength and integrity.
c. III: Desmin, GFAP, Vimentin. Muscle, glial cells, mesenchymal cells. Sarcomere
organization, integrity.
d. IV: Neurofilaments (NFL, NFM, and NFH). Neurons. Axon organization.
e. V: Lamins. Nucleus. Nuclear structure and organization.
2. IFs are NOT dynamic. Do NOT require ATP. Phosphorylation controls subunit exchange.
3. Mechanical stability. Form a network extending from the nucleus to the PM. IFs also bind
to desmosomes and hemidesmosomes. Epidermolysis bullosa simplex (involving
keratins). Amylotrophic Lateral Sclerosis (Lou Gehrigs) affects various neurofilaments.
4. Can characterize a tumors origin by the IFs it produces.