Reading...
Play button
Play button
Use LEFT and RIGHT arrow keys to navigate between flashcards;
Use UP and DOWN arrow keys to flip the card;
H to show hint;
A reads text to speech;
55 Cards in this Set
- Front
- Back
|
Which end of actin is attached to a membrane?
|
The positive (+) end
|
|
|
What 3 molecules bundle "tight bundles" of actin?
|
- fascin (which acts as spacers between filaments) or fibroblasts
- vilin - fimbrin (which also acts as a spacer) |
|
|
What molecule bundles "loose bundles" of actin?
|
alpha-actinin which acts as spacers between the actin polymers
|
|
|
Which type of bundles leaves room for the insertion of a myosin head?
|
Loose bundles, which is why they are referred to as "contractile bundles"
|
|
|
What type of actin bundle are microvilli?
|
Tight bundle
|
|
|
What type of actin bundle are stress fibers?
|
Loose bundle
|
|
|
What type of actin bundle are filapodia and lamellopodia?
|
Tight bundle
|
|
|
What type of actin bundle is responsible for the contractile ring in mitosis?
|
Loose bundle
|
|
|
Describe the function of fibronectin.
|
It is an ECM molecule. It is a homodimer (two identical polypeptide chains) in which the two chains are linked by a pair of disulfide bond bridges. Cellular integrins recognize fibronectin by its RGD sequence. If cells are placed on a Petri dish that is covered in fibronectin, the cell will flatten out to cover the dish. This "anchoring" is required for growth. Integrins in the cell’s plasma membrane bind to the fibronectin, and then oligomerize to form multiple structures called adhesion plaques, thereby permitting the cell to flatten out and move. Cancer cells do not have to flatten out to grow; they can grow in a suspension. There are multiple forms of fibronectin due to alternative splicing and they are essential for migration and differentiation in embryogenesis. It is also important in wound healing because they promote blood clotting and facilitate the migration of macrophages, they help attach cells to the ECM by binding to other ECM components and they also influence the shape of cells and the ECM.
|
|
|
Describe filamin and gelsolin.
|
Filamin cross-links f-actin which leads to gel formation, whereas gelsolin leads to the sol formation. The gelsolin in the cytoplasm is inactive when there is no calcium around, but when calcium is added, it causes the gelsolin to severe the f-actin and gelsolin binds to the ends like a cap. This causes the solution to become more viscous and less gel-like. Antibodies activate the conversion of the gel state to the sol state by binding a receptor on the surface of the cell. When the Ab is bound, the peripheral cytoplasm starts to break down in that area. Fluid in the cell rushes to that area, distorting the shape of the cell into a pseudopod. Once the pseudopod has reached it full extention, the actin network is reassembled and this causes fixation of the cell in its new position.
|
|
|
What is dystrophin?
|
A cytosolic adaptor protein made of “spring-like” spectrin repeats that binds to actin filaments and to an adhesion receptor called dystroglycan. It contains O-linked oligosaccharides that bind to various basal lamina components. Without dystrophin, the plasma membrane tears easily. Mutations reduce acetylcholine receptors on muscle cells, this leads to weakening of the mechanical stability of muscle cells as they undergo contraction and relaxation resulting in deterioration of the cells which is muscular dystrophy.
|
|
|
What protein in found on the undersurface of RBC's surface membrane?
|
Spectrin, it binds glycophorin and allows flexibility of the cell.
|
|
|
How does actin polymerization kinetics change in the presence of a nuclei core as opposed to without one?
|
When ATP is added to solution in the presence of a nuclei core, polymerization occurs instantly. Without one, it has a lag time because it must create a nuclei core before rapid polymerization can occur. This makes nucleation the rate limiting step.
|
|
|
Drescribe treadmilling.
|
It can happen whenever one end (the minus end, usually) is depolymerizing while the other end (plus end) is elongating. In the special case where the rate of growth of the plus end of the polymer equals the rate of depolymerization at the minus end, such that there is no change in the overall length of the filament, we speak of ‘steady state’ treadmilling.
|
|
|
Describe 2 well-known drugs that affect actin filaments.
|
- Cytochalasin D- it is a fungal alkaloid that depolymerizes actin by binding to the positive ends thereby "poisoning" them (inhibition of adding more actin)
- Phalloidin- it has the opposite effect; it hyperstabilizes actin filaments which drive the equilibrium in the cell such that all actin is polymerized. It binds f-actin so fluorescently-labeled phalloidin is widely used to stain for light microscopy. Poison mushrooms contain this. |
|
|
Actual measurements of actin the cell show that 40% is not polymerized. What regulated this?
|
It is closely regulated by thymosin β4 and profilin, and the nucleation process is regulated by Rho family G-proteins. Thymosin inhibits polymerization by binding to the ATP binding cleft on g-actin. When profilin binds g-actin, it promotes polymerization by allowing ADP-ATP exchange with the help of proline-rich proteins such as wasp (Wiscott-Aldrich syndrome protein) which binds to the nucleating protein arp (actin-related protein).
|
|
|
Describe cell movement.
|
- extension of one or more lamellipodia by polymerization of actin at the positive end by profilin while cofilin induces the loss of subunits from the positive end
- lamellipodia adhere to the fibronectin surface with integrins on its membrane - the bulk of the cytoplasm flows forward - the trailing edge remains attached until the tail eventually detaches and retracts |
|
|
What cellular organizations of actin filament polymerization do cdc42, Rac, and Rho induce?
|
- cdc42- filopodia, by way of activating WASP/Arp2/3
- Rac- lamellipodia by way of activating WAVE/Arp2/3 - Rho- stress fibers by way of activating Formin |
|
|
Describe the cellular distribution of microtubules.
|
They radiate out from the centrosome to fill the entire cytosol, in mitotic cells, the cytosolic array reorganizes to form the mitotic spindle apparatus.
|
|
|
Describe the structure of microtubules.
|
They are made from α- and β-tubulin which polymerizes to make protofilaments, 13 protofilaments align side to side to form a microtubule. They have (+) ends are facing away from the cell and the (-) ends are embedded in the centrosome. The α-tubulin binds GTP (the (-) end), whereas β-tubulin binds GDP (the (+) end)
|
|
|
What types of structures are made from microtubules?
|
Axons and flagella
|
|
|
Which end of a microtubule will fray?
|
The (-) end often will because it is usually composed of subunits in the GDP-form which are side-to-side “less sticky”, whereas the (+) end, which usually has subunits in the GTP form (a “GTP cap”), is “sticky”. But when the (+) end loses its GTP cap, it too will fray. But inside the cell, the minus end is usually capped, and therefore stabilized; the (+) end exhibits dynamic instability.
|
|
|
What is the main difference between actin and microtubules as far as what nucleotides they bind?
|
- actin binds ATP
- microtubules bind GTP |
|
|
During mitosis, what does the positive end of a microtubule bind to?
|
The kinetochore - it will "munch" its way back to the centrosome which will pull the chromosomes apart during anaphase.
|
|
|
How does temperature effect microtubule polymerization?
|
Higher temps make them polymerize and low temps promote depolymerization because the hydrophobic interactions between tubulin increase the entropy of the system.
|
|
|
What are 2 well-known drugs that affect microtubules?
|
- colchicines- it destabilizes them and stops mitosis
- taxol- it has the opposite effect, it hyper-stabilizes them causing all the tubulin to polymerize, this is just as bad as colchicines, taxol is a powerful anti-cancer drug |
|
|
How is the nucleate seed formed for formation of microtubules?
|
The α- and β-tubulin dimers bind to the γ-TuRC (γ-tubulin ring complex) and are stabilized by interaction with γ-tubulins. These assembled tubulin molecules act as a seen to nucleate further microtubule assembly, which determines the protofilament number.
|
|
|
What is one way to regulate dynamic instability of microtubules?
|
Adding a cap of GTP-containing subunits on the less stable region (region of GDP-containing subunits) of the MT, this will allow for growth. If the rate of addition of GTP-containing subunits slows, the ongoing rate of GTP-hydrolysis will catch up to the positive end. There will no longer be a cap of GTP-containing subunits; the protofilaments will fray and rapid depolymerization (catastrophic collapse) will begin at the plus end, until another cap of GTP-containing subunits can be formed at the plus end.
|
|
|
Describe the 2 groups of Microtubule-Associated Proteins (MAPs).
|
- one group stabilizes MTs (ex: MAP 1, 2, 4 Tau)
- one group destabilizes MTs (ex: katanin, Op18, or "stathmin") |
|
|
Describe dynein and kinesin proteins.
|
- dynein- MT-based motility such as flagella and cilia, dynein motors move toward the negative end
- kinesin- MT-based motility such as axonal transport, kinesin motors move toward the positive end |
|
|
What do neurotransmitter vesicles use to travel from the cell body all the way out to the distal synaptic membrane?
|
Kinesins and myosins
|
|
|
What do MTs govern the location of?
|
The golgi and direct many aspects of intracellular vesicle transport
|
|
|
How do vesicles travel in the cell?
|
The golgi membranes have dynein on their surface, which transports them to toward the (-) ends of MTs, so the golgi is located over the centrosome. If MTs are depolymerized, the golgi fragments into small vesicles that disperse throughout the cytoplasm. The membranes of the ER have kinesis motors on their surface, which walk along MTs towards their (+) ends, and thereby cause the ER tubules to align with MTs, and extend out towards the periphery of the cell. Again, if MTs are depolymerized, the ER collapses into the center of the cell. Like golgi, endocytic vesicles travel along MTs using dynein motors.
|
|
|
Describe intermediate filaments.
|
- the fundamental subunit is the “anti-parallel tetramer”
- they are highly diverse, ie. there are more than 20 different types which can be useful in the diagnosis of carcinomas - they are relatively stable, but can be disassembled by phosphorylation, and they are in relatively slow equilibrium with a small pool of subunits - they provide mechanical stability for the cell and strengthen cell assemblies (ie. epidermolysis bullosa simplex |
|
|
Describe epidermolysis bullosa simplex.
|
Mutant keratin does not properly assemble into IFs, therefore epidermal cells are not mechanically tought, and tear in response to small shear forces; consequently, epidermal layer of skins separates from underlying dermis, forming blisters.
|
|
|
Most intermediate filaments have what in common?
|
A central, α-helical, rodlike domain, approx 300 aa long and the amino-terminal and carboxy-terminal domains are non-α-helical and vary greatly in size and sequence among different IF subunit proteins. These wrap around each other to form an anti-parallel tetramer that do not have (+) or (-) ends; they are symmetric.
|
|
|
What is a benefit of having many different isotypes of IFs?
|
A specific Ab against a keratin (a type of IF) expressing in a particular cell type can be used as a sensitive way to detect rare metastatic cancerous liver cells in nearby sentinel lymph nodes. None of the other cell types found in lymph nodes will express the particular keratin found in liver cells.
|
|
|
Describe the nuclear lamina and what happens to it in mitosis.
|
- made from IFs
- lies on the inner surface of the inner membrane of the double membrane nuclear envelope - at the onset of mitosis, cyclin-dependent kinase phosphorylates nuclear lamins, causing them to disassemble. This permits the nuclear envelope to break up into vesicles, and frees the chromosomes to bind spindle fibers. At the end of mitosis, a phosphatase removes the phosphate from the lamins and they reassemble, leading to the reassembly of the nuclear envelope. |
|
|
Describe the cell cycle.
|
- G1 is the synthesis of RNAs and proteins; there are 2n chromosomes, hours 0-5
- S is the synthesis of DNA and chromosome replication; 2nà 4n; hours 5-12 - G2 contains 4n chromosomes; hours 12-15 - M phase is hours 15-16 |
|
|
Describe the G0 phase.
|
It is a state in which animal cells are very metabolically active with high rates of protein synthesis, its just that cells are in a steady state, where the overall rate of cellular protein synthesis is balanced by equal rate of overall protein degradation; most cells spend their entire life in the G0 state; growth hormones stimulate cells to leave G0 and enter G1.
|
|
|
Describe the G1 restriction point.
|
Consider G-0 cells that have been stimulated to leave G0 by the addition of growth factors to the culture medium. For the first few hours after addition of growth factors, it is the case that, if the growth factors are removed, the treated cells will return to G0. But there comes a time, called the restriction point, where the cells have reached a point of no return. If growth factors are removed after this time, they will not return to G0; instead, they will initiated DNA synthesis a few hours later and progress through to mitosis.
|
|
|
What does the cytoplasm of a dividing cell contain?
|
A “factor” that causes other cells to enter M phase also if that cell is fused with a nondividing cell. This Mitosis Promoting Factor (MPF) is dominant over G1, S or G2. It was discovered from an experiment with frog oocytes. Cytoplasm from a mature cell was injected into an immature cell and that cell was induced into maturation, just like progesterone does.
|
|
|
Describe in more detail MPF.
|
It is a serine kinase. It contains 2 subunits: a regulatory subunit called cyclin and a catalytic or kinase subunit called cyclin-dependent kinase (cdk). The function of cyclin is that the binding of cyclin leads to activation of the kinase subunit and that cyclin determines substrate specificity of the kinase subunit (ie. which proteins will be recognized and phosphorylated). The function of cdk is to phosphorylate various proteins which causes the initiation of the “mitosis program” (starts mitosis) and that one of the proteins that is phosphorylated by cdk in turn causes the destruction of cyclin; this results in the inactivation of cdk and the termination of the “mitosis program” (ends mitosis).
|
|
|
In which phase of the cell cycle is the concentration of mitotic cyclin the highest? How does this compare to the rate of cyclin synthesis?
|
Mitosis. The rate of mitotic cyclin (cyclin B) synthesis remains constant throughout the cell cycle in fertilized sea urchin eggs, but the rate of proteolysis varies, increasing and decreasing the amount of cyclin that is present in the cell.
|
|
|
Describe cdc2+, cdc 2- and cdc2d cells.
|
- cdc2+ is a normal cell that makes two normal sized daughter cells
- cdc2- is a cell that cannot enter mitosis so the cells grows extremely large - cdc2d cells enter mitosis too early, making two very small “wee” cells |
|
|
Name 5 substrates of MPF.
|
- anaphase-promoting complex : phosphorylated, activation of ability to bind to cyclin, initiating the ubiquitination and subsequent proteolysis of cyclin. This causes the inactivation of MPF, which is required for the late events of M phase. Activated APC also targets a protein called securing for ubiquitination and destruction, leading to permitting sister chromotids to separate during anaphase
- nuclear lamins: phosphorylation causes disassembly of the lamina, which is part of the breakdown of the nuclear envelope during mitosis - condensin: phosphorylation activates the condensation of chromosomes during mitosis - Microtubule-Associated Proteins: phosphorylation of these is involved in the reorganization of MTs to form the mitotic spindle - myosin II light chain: phosphorylation at an inhibitory site by MPF inhibits the activation of the myosin by MLC kinase; subsequent dephosphorylation of this inhibitory phosphate leads to cytokinesis |
|
|
Describe the connections between MPF, APC, cyclin and the process of ubiquitination-mediated proteolysis.
|
APC is an E3 Ubiquitin ligase; when it is phosphorylated by MPF, it is activated and binds to the “destruction box” sequence on cyclin B (the most common form of mitotic cyclin). It then recruits ubiquitin proteins from E2 (a ubiquitin carrier) and transfers them (the ubiquitin molecules) onto an adjacent lysine (the lysine side chain amino group) of the cyclin B molecule. After 4-6 ub’s are added, it is now tagged to go to the proteosome. As it enters the proteosome, the individual ubiquitin molecules are cleaved off and recycled for subsequent use; the cyclin protein is pushed into the internal chamber and cleaved into small peptide fragments by the proteases that make up the walls of this cylindrical complex.
|
|
|
Summarize the roles of MPF and APC in the cell cycle as discussed in lecture
|
(1) MPF initiates anaphase by phosphorylating APCcdc20
- phosphorylated (thereby activated) APCcdc20 binds a destruction box sequence on Securin. - polyubiquitination/proteolysis of securin occurs, permitting separase to cleave the ‘glue’ holding the metaphase chromatid pairs together. - this ends metaphase and initiates anaphase. (Thereafter, APCcdc20 releases its cdc20 partner and (temporarily) returns to an inactive state. Later, APC complexes acquires the cdh1 partner, but still stay inactive, until the next step): (2) MPF inactivates itself after telophase, thereby permitting to onset of telophase and cytokinesis. It does this by phosphorylating APCcdh1. - phosphorylated (thereby activated) APCcdh1 binds a destruction box sequence on cyclin. - polyubiquitination/proteolysis of cyclin occurs, thereby ending the catalytic activity of its cdk partner; ie, MPF activity ceases. - The inactivation of MPF permits the onset of telophase and cytokinesis. |
|
|
How are sister chromatids separated during anaphase?
|
- securin, which inhibits separase, is polyubiquinated by APC. This leads to the destruction of securin.
- this releases separase from inhibition, which now cleaves the cohesion between the two |
|
|
What is condensin?
|
A protein that uses ATP to condense chromosomes during mitosis.
|
|
|
Describe Rb.
|
Gets its name from retinoblastoma, a hereditary tumor that develops early in life; it is a tumor repressor gene and having one good copy of the gene can prevent cancer. The mechanism behind this is that Rb binds (and thereby inhibits) a molecule of the transcription factor E2F. Sequential phosphorylation of Rb by cyclin D/cdk4 and cyclin E/cdk2 causes Rb to release E2F. When it is free, E2F molecules activate transcription of the genes encoding enzymes for the initiation of DNA synthesis. Normally, in resting G0 cells, E2F is bound to Rb, and therefore not able to transcribe the genes for DNA synthesis.
|
|
|
How does acetylation affect histones?
|
- acetylation opens up DNA for activation
- deacetylation makes DNA inaccessible – a “repressed” region of a chromosome is deacet. |
|
|
Describe the molecular basis of the restriction point in G1.
|
Normally, cyclin D is very unstable, but in the presence of hormones, it is protected from degradation. Cyclin D-cdk4 and cyclin E-cdk2 act sequentially to hyperphosphorylate Rb and release Rb from E2F to start DNA synthesis.
|
|
|
What happens if DNA damage is detected?
|
If damage is detected in G1, S or G2, the cell arrests in the cell cycle until the damage is repaired, if the damage is too extensive, apoptosis is initiated. The DNA damage checkpoint act through p53. The amount of p53 (a transcription factor) rises when damage is detected. If there are mutations in p53, it can no longer “tumor suppress”. The level of p53 that is synthesized is very high but the rate of degradation is also very high. This is so that if damage is detected, the amount of p53 can rise dramatically by just shutting off its degradation.
|
|
|
What is another name for p21cip and what does it do?
|
The universal inhibitor gene, the cip stands for cyclin inhibitor protein. This is activated by high levels of p53 (the transcription factor) so that entry into the S phase is blocked.
|
