Lecture 5 - Cytoskeleton Flashcards
What is the diameter of microtubules?
24-25nm
What is the diameter of microfilaments?
6-7nm
What is the general organization of microtubules?
Curvy structures that radiate from mitochondrial organizing center (MTOC)
Where is the general organization/location of microfilaments?
Linear bundles beneath the plasma membrane
What is the diameter of intermediate filaments?
10nm
What are the subunits of microtubules?
Tubulin heterodimer, each made up of alpha and beta tubulin
What molecule do all tubulin monomers bind?
GTP
What provides the energy for polymerization of microtubules?
GTP
Microtubules have a (+) and (-). What is the significance of these ends?
(+) end is bound to GTP and is the side that favors additional polymerization. (-) is bound to GDP and this favors dissociation.
What is the primary function of microtubules?
Transport (organelles within cell, along axon)
What end to kinesins move towards?
(+) end: TureKKKK is (+) about the state of the earth.
What end to dyneins move towards?
(-) end: Dying is (-).
By what mechanism do vinblastine, vincristine, nocodazole, and colchicine inhibit cell division?
Prevent assembly (polymerization) at (+) end of microtubules.
By what mechanism does paclitaxel (Taxol) inhibit cell division?
Taxol binds (-) end and prevent the dynamic assembly and disassembly of microtubules required during cellular division.
What is the role of a basal body?
Located at the base of a cilia and organizes an array of microtubules.
What is the pattern of microtubule branching in basal bodies?
9 x 3 (with one microtubule in the middle of the 9 groups)
How many monomers long is a complete turn of a standard microfilament?
14 (37nm)
What is the name of monomeric actin?
G-actin (G for globular)
What is the name of polymerized actin?
F-actin (F for filamentous)
What molecule promotes polymerization of G-actin?
ATP
Which is more stable: microfilaments or microtubules?
Microfilaments
What is “treadmilling?”
Continuous growth and (+) end at same rate of continuous disassembly at (-) end.
What proteins (3) regulate actin polymerization and what is the role in regulation of each protein?
1) Profilin - promotes actin assembly on growing F-actin filaments, binds g-actin and prevents polymerization, functions in ADP to ATP exchange in actin monomers to to prepare for polymerization into F-actin
2) Thymosin - negative regulation by binding g-actin
3) Gelsolin - sever & cap actin filaments, capping prevents addition and loss of actin monomers. Functions in response to Ca2+, pH, and phospholipids.
Where are actin stress fibers located?
Bundled along basal cell surface and terminate at focal adhesions.
What organizes the actin of stress fibers?
alpha-actinin & talin
Where are actin adhesion belts located?
Circumferential actin near apical cell surface
What is the role of actin adhesion belts?
Stabilize cadherin-mediated adherens junctions
What organizes actin adhesion belts?
catenins & alpha-actinin
What actin structure provides a scaffold for intracellular signal transduction?
Adhesion belts
A fungal toxin (and alkaloid) that binds (+) end of actin and blocks polymerization.
Cytochalasins
Amanita mushroom poison, binds f-actin and prevents disassembly of (-) end
Phalloidin
What is the remedy for phalloidin poisoning?
Raw hamburger
Bind G-actin and induces F-actin depolymerization (derived from sea sponge)
Latrunculins
Fingerlike extension at cell surface that increase surface area
Microvilli
What organizes the actin of microvilli?
villin and fimbrin
What causes hereditary spherocytosis?
mutant spectrin
What protein cross-links actin into 2D mesh?
spectrin
Where is spectrin most prominent?
RBCs (erythrocytes), but ubiquitous
What links spectrin to Band 3 and Glycophorin?
Ankyrin
What regulates the assembly and disassembly of intermediate filaments?
phosphorylation
What are the six types of intermediate filaments?
1) Acidic keratins
2) Basic to neutral keratins
3) Vimentin, Desmin, Glial fibrillary acid protein (GFAP), Peripherin
4) alpha-internexin
5) Nuclear laminins
6) Nestin
What is the significance of cytokeratins?
Change in cytokeratin expression is among the most common markers of neoplasia. All epithelial cells express at least on AK and one B-NK.
List the Type III intermediate filaments and their roles/significance.
Vimentin - mesenchymal, endothelial, and leukocytes. Useful marker of dedifferentiation of invasive cancer carcinomas.
Desmin - skeletal muscle in Z-disk, smooth muscle
Glial fibrillary acid protein (GFAP) - astrocytes and Schwann cells
Peripherin - neurons of peripheral nervous system
(VDGP)
What type of intermediate filaments run along axons and dendrites and can be greater than 1 meter in length?
Type IV
Where is alpha-internexin found and what type of intermediate filament is it?
CNS-spinal cord, optic nerve; Type IV
What type of intermediate filament are nuclear laminins?
Type V
1) What is significant about nuclear laminins, 2) what structure do they support, 3) and what can cause rapid disassembly?
1) FORM INTO A SQUARE LATTICE (All other intermediate filaments form fibers.) 2) Nuclear membrane 3) Serine phosphorylation
What type of intermediate filament is Nestin and where is Nestin found?
Type VI, Stem cells of CNS
What is the approximate size of a mitochondrion?
0.2 uM to 7uM
What structure in the mitochondrial outer membrane allows molecules
Porins
What tissues/organs are most affected by mitochondrial diseases?
High energy organs/tissues
MITOCHONDRIAL DISEASE: Paralysis of eye muscles & mitochondrial myopathy
CPEO (chronic progressive externa opthalmoplegia)
MITOCHONDRIAL DISEASE: Abnormal movements involving muscular rigidity; frequently accompanied by degeneration of the basal ganglia of the brain.
Dystonia
MITOCHONDRIAL DISEADE: CPEO combined with such disorders as retinal deterioration, heart disease, hearing loss, diabetes, & kidney failure
KSS (Kearns-Sayre syndrome)
MITOCHONDRIAL DISEASE: Progressive loss of motor & verbal skills and degeneration of the basal ganglia; potentially lethal childhood disease.
Leigh’s syndrome
MITOCHONDRIAL DISEASE: Permanent or temporary blindness stemming from damage to the optic nerve.
LHON (Leber’s hereditary optic neuropathy)
What effect does mitochondrial dysfunction have on sperm?
Immotile - leading to male infertility
In an H&E stain how does rER stain?
Purple with hematoxylin (basophilic)
What does an extensive rER indicate when seen in a cell?
That cell likely synthesizes many proteins for secretion
Where is cholesterol produced?
ER
Where is ceramide produced and what is makes up ceramide?
ER, serine + FA = sphingosine & sphingosine + FA = ceramide
What is ceramide a precursor for?
Glycolipids and sphingomyelin
Where are N-linked (Asn) oligosaccharides often attached to proteins?
ER
Where does steroid and triglyceride synthesis occur?
sER
What organelle is involved in detoxification?
sER
What are the major functions (5) of the golgi?
1) Modify N-oligosaccharides 2) Distribute non-cytosolic proteins to plasma membrane, lysosomes, & secretory vesicles. 3) Assembles proteoglycans from proteoglycan core proteins 4) Adds mannose-6-phosphate to proteins that are targeted for lysosomes 5) Sulfates sugars on proteoglycans.
Where will a protein tagged with mannose-6-phosphate in the golgi end up?
A lysosome
Where are peroxisomes primarily found?
LIVER, kidney pct, skeletal muscle
What are the primary oxidative enzymes (3) found in peroxisomes?
1) D-amino acid oxidate
2) urate oxidase
3) catalase
What is the diameter of a peroxisome?
0.15 - 0.25 uM
What are the functions of peroxisomes in human liver specifically?
1) Oxidize excess FAs
2) Breakdown H2O2 (is a product of FA ox, broken down by catalase)
3) Cholesterol synthesis
4) Synthesis of bile acids
5) Synthesis of lipids used to make myelin
6) Breakdown of excessive purines (AMP, GMP) to uric acid
Lethal condition (1st decade of life) where no functional peroxisomes are produced. Peroxisomal enzymes are not transported to organelle (peroxisome membranes remain empty).
Zellweger syndrome
What causes Zellweger syndrome?
Genetic defect: gene encoding the receptor for peroxisome targeted proteins is defective.
Is the plasma membrane visible with LM?
No - need TEM
Is the rER visible with LM?
Yes - stains basophilic (purple/blue)
Is the sER visible with LM?
Yes - when abundant, clear “moth-eaten” appearance to cell
Is the golgi visible with LM?
Yes - clear region near nucleus (ex. plasma cell)
Is the mitochondria visible with LM?
Yes - cytoplasm appears granular, stain with Janus green B
Are lysosomes visible with LM?
Yes - stain for acid phosphatase
Are endosomes visible with LM?
Yes - only large ones, still difficult to see
Are peroxisomes visible with LM?
Yes - stain for catalase
Are microtubles visible with LM?
Yes - only “bundles” that are stained with fluorescent dye or use of phase contrast imaging
Are centrioles visible with LM?
Yes - with fluorescent tag
Are intermediate filaments visible with LM?
Yes - only bundles, not individual filaments
Are ribosomes visible with LM?
No
Are microfilaments visible with LM?
Yes - bundles with fluorescent dyes
What protein transports vesicles between ER & golgi?
COP-II (Coat protein II)
What protein transports vesicles between golgi stacks?
COP-I (Coat protein I)
What protein mediates vesicle transport between the plasma membrane and the TGN?
Clathrin
What is the primary and secondary function of lysosomes?
Primary - storage Secondary - degradative processes
How do lysosomes become acidified?
Proton pump in membrane
What coat protein facilitates pinocytosis?
caveolin
Ingestion and degradation of foreign material take into the cell by receptor-mediated endocytosis or phagocytosis
Heterophagy
Segregation and digestion of an organelle or other cell component with membranes from the rER.
Autophagy
What protein facilitates receptor-mediated endocytosis?
Clathrin
How many clathrin coated pits exist per cell and what percent of the cell surface is this. What percent of cell receptors are found in coated pits?
500-1000 pits/cell, 2-4% of cell surface, 70% of cellular receptors
What are the three possible outcomes of receptor-mediated endocytosis (provide an example for each)?
1) Ligand degraded in lysosome and receptor recycled (ex LDL)
2) Ligand and receptor recycled (ex transferrin)
3) Ligand and receptor both degraded (Epidermal growth factor)
How do cholera and tetanus toxin get into cells?
Clathrin independent pathway
Deficiency in hexosaminidase resulting in accumulation of glycolipids (gangliosides) in NS.
Tay-Sachs
Defective glucocerebrosidase causing glucocerebrosides to accumulate in the CNS and spleen.
Gaucher’s syndrome
Defective sphingomyelinase causing accumulation of sphingomyelin and cholesterol in CNS and spleen.
Neiman-Pick disease
Low occurance of LDL receptor, binds LDL poorly or LDL cannot be internalized.
Hypercholesterolemia
How does Ebstein-Barr Virus (EBV) enter cells? What does EBC cause?
Binds to receptor present on B cells. Causes mononucleosis & contributes to development of Burkitts lymphoma
How does Influenza virus enter cells?
Hemagglutinin binds to carbohydrates on target cell surface and is endocytosed.
Failure to “tag” (by phosphorylation) all the hydrolytic enzymes that are supposed to be transported from the golgi to the lysosomes (they lack M6P tag).
I-cell disease (“inclusion cell disease)
Failure to synthesize an alpha-L-iduronidase.
Mucopolysaccharidosis (MPS-1)
What is the function of alpha-L-iduronidase?
It is required to break down proteoglycans like heparan sulfate.
What enzyme is the function of Beta-N-acetylhexosaminidase? (Also, what disease results from its deficiency?)
Converts ceramide to to glucose.
What are the hallmarks of LSD?
- Unusual fascial features
- Cloudy eyes
- Purplish/bluish skin
- Distended belly
- Short stature
- Muscle weakness/lack of muscle control
What causes sickle cell anemia?
Modification of spectrin skeleton
