Cytoskeleton, microtubules Flashcards
Describe microtubules in general.
- tubulin subunits
- hollow - provides rigidity and strength
- polar
- dynamic - population of tubules and subunits constantly exchanging
- highly conserved among organisms, meaning it is super important
- railways for motor proteins - kinesin, dynein which walk by hydrolyzing ATP and directed by polarity
Describe actin filaments in general.
very similar to microtubules
- made from actin subunits
- non-hollow
- polar
- dynamic
- highly conserved
- railways for motor proteins
Describe intermediate filaments in general.
- not very conserved
- various types of cell-specific intermediate protein subunits
- non-hollow
- non-polar
- non-dynamic - not many free subunits available in the cell
- diverse
- not railways for motor proteins
- less essential to the cell than microtubules and actin. Only become very important if one of the other two types become defective.
How are intermediate filaments used as markers of disease?
Because they are cell-type and location specific.
What is an experimental method that is useful for studying functions of microtubule-associated proteins?
RNA interference
function-blocking antibodies
Is GTP hydrolysis required for microtubule assembly?
Hydrolysis is not required, but GTP attached to tubulin is required.
How does free tubulin exist?
As GTP-tubulin
What is the polarity of microtubules in neurons?
The plus end of microtubules is oriented away from the cell body (towards end of axon)
How can microtubules and actin be visualized together?
Cells can be made to express fluorescent hybrids of the subunits that fluoresce at different wavelengths. Images can then be merged to view the structures simultaneously.
How is it that cytoskeletal elements sometimes overlap in function?
because both actin and myosin play roles in coordinating the cell and dividing labor.
Describe the subunits of microtubules.
- subunits exist in as a dimer of alpha and beta tubulin
- cells contain both microtubules and free tubulin dimers, which are constantly in exchange
- each ring of the microtubule consists of 13 subunits around
Describe the polarity of microtubules and how polarity is established.
- polarity has nothing to do with chemical charge!
- plus and minus end because subunits are heterodimers
- plus end is favored for assembly and disassembly (more dynamic!)
- polarity exists all along the length of the microtubule
- alpha subunit revealed at minus end, and beta subunit is revealed at the plus end
Describe how microtubules are non-equilibrium polymers.
There is first a lag phase of microtubule polymerization, followed by an exponential growth phase, and finally a steady state where subunits are coming on and off
This is a non-equilibrium polymer because tubulin must be above a critical concentration before polymerization can take place, much like salt cannot crystallize in water until it reaches a critical concentration.
What conditions are needed for a microtubule to form?
- GTP
- 37 degrees
- magnesium
- tubulin above critical concentration
How was microtubule polarity discovered?
imaging of flagella revealed that the two microtubule ends have different critical concentrations for assembly, plus end having a lower critical concentration and the minus end having a higher critical concentration.
What is an example of an organelle which is distributed by motor proteins on microtubules?
Golgi. The motor proteins involved are highly processive.
Describe microtubule treadmilling.
Treadmilling is an illusion of microtubule movement which is observed in vitro. It is the tubulin subunit concentration is at a point in which polymerization at the plus end equals depolymerization at the minus end, so the microtubule appears to move through solution. This does not seem to happen in vivo.
Describe the growth and breakdown of microtubules
- GTP tubulin adds to plus end, generating GTP cap. As long as there is a GTP cap, microtubule will continue to assemble
- GTP hydrolysis occurs only after GTP-tubulin is incorporated into the microtubule. Allows energy to be stored in the microtubule lattice.
- If GTP hydrolysis catches up to assembly, GTP cap is lost and microtubule disassembles through catastrophe
- If GTP cap is re-established before microtubule is completely gone, we get rescue
Describe the types of molecular motor proteins, and what they are good for.
- Dynein carries retrograde cargo and travels toward the minus end. Processivity is enhanced by dynactin.
- kinesis carries anterograde cargo and travels toward the plus end
- both consist of dimers, because they move by walking. This allows them to be highly processive.
- Both are ATPases that generate forces to create motion in the cell.
- both have globular motor domains (dimerized part), cargo domain, and a stalk domain in between that are all part of the same gene product.
Why do different cell types have microtubules organized in different ways?
Different microtubule organization allows for different types of cell work to be accomplished.
Describe ciliar microtubule arrays.
- microtubule minus end is embedded in the basal body (the cell body). In the basal body, there are no free tubules at the center of the structure, and it consists of 9 +3 triplet centriole.
- the plus end is pointed outward along the cilia or flagella. Microtubules are arranged 9+2 around the circumference of the flagella, and are highly crosslinked. Two central singlet microtubules are at the center of the flagella.
- These microtubules are very stable.
- The flagella beats through flagellar dynein.
Describe the structure of the centrosome.
- pair of centrioles
- each centriole consists of microtubule triplets
- surrounded by pericentriolar matrix
- PCM contains rings of gamma tubulin which serve as nucleating sites, where microtubules are very stable
Describe gamma tubulin and how it functions.
- in pericentriolar matrix
- as different from beta and alpha tubulin as they are from one another
- present in the form of gamma turc (gamma tubulin ring complex)
- gamma turc establishes protofilament number (13) and polarity orientation of the microtubule
How are complex “non-centrosomal” microtubule arrays established?
- may be multiple centrosomes in the cell not known
- gamma turc may be distibuted to non-centrosome locations in the cell (can be determined from treatment with nocodazole. Microtubules are destroyed, and where new ones appear tells us where gamma turc is located in the cell)
- microtubules may break off from centrosome and move around the cell in a new pattern
Describe microtubule movement in living cells.
Microtubule movement is entirely relative. If the cargo which is carried along it cannot move because it is attached to a coverslip, the microtubule itself will move along it.
How is axon branch fomation accomplished in neurons?
Axon branch formation is accomplished through microtubule cut and run, via two distinct methods:
- katanin mode: widespread, tau makes microtubules more vulnerable to katanin
- spastin mode: spastin is recruited specifically to the branch location
