Microtubules Flashcards
Functiosn of mt’s
Make up the mitotic spindle
Arranging cellular organelles during interphase
Serving as tracks for celllular constituents to be transported
Cilia & flagellar movement
Taxol as cancer therapy
Stabilizes mt’s so that cells can’t perform mitosis (no anaphase)
The end ringed by _tubulin is the negative end
The end ringed by _tubulin is the positive end
Which holds the ATP that gets hydrolyzed?
a-tubulin -> negative
B-tubulin -> positive ; holds the ATP that gest hydrolyzed
MTs fom 3 distinct ring structures- what are their functions?
Single ring- interphase & mitosis
Doublet - cilia & flagella
Triplet- basal bodies and centrioles involved in mt organizing centers (MTOCs)
Explain this graph
MT assembly depends on dimer pool concentration
At low [monomer], dimers form spontaneously.
As [monomer] increases, a critical conc (Cc) is reached; at or above the Cc, dimers spontaneously polymerize into MTs and dimer conc no longer increases.
Why does assembly and disassembly occu rmore rapidly at the (+) end than the (-) end?
The (+) end has a lower critical concentration - less dimers are needed to stimulate polymerization from the (+) end
The B tubulin added to the (+) end must
be GTP-bound
Catastrophe- what is it and how do you rescue?
Catastrophe: rapid depolymerization of the (+) end because the GTP-bound dimer pool is far below Cc(+).
Simply increase [GTP-dimer]
When does treadmilling occur? What is it?
Treadmilling: dimers are added to the (+) end but removed at the (-) end
[dimer] is greater than Cc(+) but less than Cc(-)
What happens as more MTs are polymerized and [GTP-dimer] drops?
Assembly slows until [GTP-dimer] is less than Cc(+) –> GTP cap is hydrolyzed, producing an unstabel GDP cap and catastrophe
MT associated proteins (MAPs)
Stabilize the MT because
- they contain positive amino acids in their MAP basic domain that bind to negatively charged amino acids in tubulin –> reduces charge repulsion between neighboring tubulin subunits
- They have a negative acidic projection domain that maintains the distance between MTs
Tau, MAP2, MAP4
How do you inactivate MAPs to destabilize to MTs?
Phosphorylation of their positive basic domain to create negative charges
MT destabilizing proteins: Kinesin 13, Stathmin, Katanin
Kinesin-13: Family of proteins that binds to the end of the MT and bends the protofilaments to promote catastrophe and
Stathmin/OP18: also binds to the end and bends, but removes two dimers at once; may also promote GTP hydrolysis; inactivated by phosphorylation
Katanin: sever or induce breaks in the MTs to expose new GDP-caps that promote catastrophe at the newly formed (+) end
Colchicine
Binds and sequesters tubulin dimers to bring [GTP-dimer] below Cc(+), preventing polymerization and promoting catastrophe.
Used to reduce WBC migration and inflammation in gout
Vinblastine , Podophyllotoxin, and Nocodazole
cancer treatment by inhibiting mitosis (like taxol)
Cells are arrested in G2 (Nocdazole used in the lab to synchronize cells in culture)
Alzheimer’s disease and Tau
Tau is a MAP that stabilizes MTs.
In AD, it is hyperphosphorylated and dissociates from the MT –> forms neurofibrillary tangles that lead to neuronal cell death
Significance of a phosphothreonine-proline motif in Tau and APP
If the proline is in a cis-configuration, then phosphorylation is accelerated and promotes the formation of amyloid plaques from APP and neurofibrillary tangles from Tau
Pin1
Converts spontaneously formed cis-prolines in phosphothreonine-proline back into trans -
-> dephosphorylation of both APP and Tau
–> prevents deposition of amyliod plaques and neurofibrillary tangles
MT organizing centers- what does it do and what are otehr names?
Nucleate the initial polymerization of MTs; has the (-) pole embedded and stabilized in its matrix
Centrosome in interphase when it helps organize the spatial arrangement of organelles
Spindle poles when organizing the mitotic spindles
MTOC for the mts of axons
Basal bodies for cilia and flagella
MTOCs either possess or are derived from
Centrioles: triplet-ringed MT structures
MTs transport vesicles and organelles.
How is bidirectional cargo transport possible?
-
Kinesin motor protein binds cargo and B-tubulin subunits –> walk across subunits to transport cargo toward the (+) end
- Anterograde
- Uses ATP hydrolysis
-
Dynein motor protein: retrograde toward (-) end
- Interact with dynactin, which binds the cargo
- Head domain provides ATPase fxn; hydrolysis moves the head with respect to the stem
- Stalk domain binds MT and moves when ATP hydrolysis causes the power stroke
When do MTs use their track funciton?
To organize organelles in teh cell
To move vesicles (e.g. secretory vesicles to the cell surfacE)
In exocytosis of a secretory vesicle, the secretory vesicle is brought to the cell surface along MTs using ___. The vesicle is then exchanged from the MT to the ___ so __ can bring the vesicle the rest ofthe way to the membrane for exocytosis.
Use kinesins to get to the actin cytoskeleton, so that myosin motors can bring teh vesicle the rest of the way
During endocytosis, molecules are brought into the cell in ____, which are transported to their final destination in teh cell along the __ using ___
Enter as endosomes
Transported to their final desitnation via MTs using dynein
What is this?
Doublet MTs –> it’s cilia and flagella
Each doublet consists of A and B tubules
The outer and inner dynein of a cilia/flagella connect to the _ tubule as cargo and then motor along the _tubule of the adjacent double toward the __pole at the base.
The dyneins connect to the A tubule as cargo and then motor along the B tubule of the adjacent doublet toward the (-) pole.
Rhythmic sliding of adjacent mt filaments due to these axonemal dyneins move the cilia or flagella
Ciliary dyskinesia
immotile ciliary syndrome arising from dynein dysfunction.
50% of people with this have a mirror-image reversal of their internal organs (situs inversus totalis) –> Kartagener syndrome
Poor mucociliary clearance, affected hearing, infertility due to fallopian tubes or sperm
MTs in early interphase
Centrosome (later called the “spindle pole”) begins to duplicate and new daughter centrioles bud from the centrosomal matrix of the MTOC.
MTs in prophase
The interphase MTs begin to degrade and get replaced by mitotic spindle MTs, all of which originate from the replicated spindle pole and separate to opposite sides of the nucleus.
First one is the Aster MTs, which separate to opposite sies o the nucleus.
Chromsomes are condensing
Prometaphase
- Nuclear membrane breaks down
- Central spindle region is established and the kinetochore & polar MTs extend toward the central spindle region
- Condensed chromsomes are captured
Metaphase
Condensed chromosomes are aligned equidistant between the centrosomes at the metaphasic plate
Anaphase
Separation of chromosomes.
- Cohesin holding the sister chromatids together at the centromere is broken down.
- Kinetochore MTs diassemble, pulling the sister chromatids to the opposing centrosomes
- Meanwhile, the centrosomes are pulled farther apart from one another.
Telophase & cytokinesis
- Nuclear membrane reforms around each diploid set of chromosomes
- Myosin contractile ring condenses
- Mitotic MTs break down and are reorganized back into the interphase MT arrangement
Three types of mitotic MTs
- Aster/astral MTs run from the spindle poles toward the cell cortex; orients the spindle poles w/ respect to the axis of cell division
- Kinetochore MTs bind the kinetochores to pull apart the chromatids
-
Polar MTs:
- push duplicated centrosomes apart in prometaphase
- maintain orientation of the central spindle
- push the spindle poles furhter apart in late anaphase
Centrosome (spindle pole) migration
Aster MTs connect the spindle poles to the cell cortex
Dyneins (green) on aster MTs attached to the cortex motor toward the (-) end, pulling the spindle poles apart.
Kinesins (yellow) on polar MTs motor toward the (+) end of the opposing polar MTs, pushing the poles apart.
Chromosome kinetochore end capture
- Kinetochore MTs bind the kinetochore
- Kinetochore-associated kinesins move along the kinetochore MT toward the (+) end until the chromosome is end-captured.
- End-captured MTs are stabilized by their interactions with the chromosome’s kinetochore.
Chromosome alignment of metaphase- how do they get aligned perfectly equidistant between the two spindle poles?
To push it forward, kinesin-7 can hold onto the kinetochore and motor toward the growing (+) end.
To pull i tback, kinesin-13 is an MT destabilizing protein; as MT depolymerizes, a dynein will pull it toward the (-) pole
Anaphase: the kinetochore MTs disassemble to drag the chromatids back toward each othe spindl epoles.
Anaphase A vs B
Anaphase A: kinetochore MTs disassemble at both (+) and (-) ends by kinesin-13
Anaphase B:
- Kinesin motors on opposing polar MTs, pushing the polar MTs and spindle poles apart.
- Sister dynein proteins attached to the cortex motor toward the (-) end of the aster MTs, pulling the spindle poles apart.
What is the structure of the tubulin dimer sin regard to the status of the GTP
MTs are repeating a-tuulin and b-tubulin monomers, which form ab-tubulin heterodimers. There’s a GTP in both, but the GTP in a-tubulin is never hydrolyzed but the GTP in the B-tubulin dimer can be hydrolyzed
At low tubulin concentrations, tubulin monomers
spontaneously assemble into ab-tubulin heterodimers.
As conc increases to critical concentration, the dimers then form into protofilaments and microtubules.
