Cytoskeleton (ch 16) Flashcards
What are the 3 types of cytoskeletal filaments?
- Intermediate filaments
- Microtubules
- Actin filaments
What is the main purpose of intermediate filaments?
Provide mechanical strength to the cell.
What are the 2 main purposes of microtubules?
- Determine organelle positioning
2. Direct intracellular transport
What are the 2 main purposes of actin filaments?
- Determine the shape of the cell surface
2. Required for whole cell locomotion
What is the most conserved protein in animal cells?
Actin.
What animal species lack intermediate filaments?
Species which have exoskeletons.
What are the relative widths of intermediate filaments, actin filaments, and microtubules?
Intermediate filaments: 10nm
Actin filaments: 5-9nm
Microtubules: 25nm
Give an example of a way in which the cytoskeleton is dynamic.
Example: rapid cytoskeletal rearrangement in neutrophils allows them to chase down bacteria.
Example actin forms a contractile ring which pinches off the cells during cytokinesis.
How are cytoskeletal filaments responsible for polarity within the cell?
They have +ve and -ve ends which direct cell components to one side of the cell or the other.
Why does the cell disassemble and reassemble filaments instead of moving assembled filaments around?
Faster, less awkward to move them through the crowded cytosol.
What energy source drives microtubule formation? What about actin filament formation?
Microtubules: GTP
Actin: ATP
Why have a microtubule made from many protofilaments?
Protofilaments alone are thermally unstable, but a microtubule composed of protofilaments is much harder to break.
How many protofilaments make up a microtubule in mammalian cells?
13.
What is characteristic of actin in its T-form? What about its D-form?
T-form: bound to ATP
D-form: bound to ADP
At what end of the microtubule or actin microfilament are new monomers most readily added?
+ve end.
Which form of actin (T or D) is more likely to be depolymerized?
The ADP bound D-form.
Is depolymerization of actin dependent on concentration? Please elaborate.
No. Subunits leave polymer end at constant rate regardless of concentration.
Is polymerization of actin dependent on concentration? Please elaborate.
Yes. The # of monomers that add to the polymer per unit time is proportional to the [free subunit].
What is the “persistence length” in relation to actin?
The maximum length at which two points along an actin filament will be oriented in the same direction. Beyond this length the filament is too bendy and the directions are uncorrelated.
What is “treadmilling” in actin filaments?
When the actin filament is gaining and losing monomers at the same rate so the length of the filament remains constant.
How can actin treadmilling propel cellular movement?
Net assembly at the leading edge pushes the cell forward while net disassembly at the lagging edge pulls up the rear of the cell.
In actin polymerization, what effect will profilin have when it binds to free actin monomers?
It will promote rapid +ve end growth.
In actin polymerization, what effect will thymosin have when it binds to free actin monomers?
It will totally inhibit growth at the +ve end of an actin filament.
Where in the cell does actin filament nucleation most frequently occur?
At or near the plasma membrane.
What 2 proteins are most closely associated with actin filament nucleation?
- Arp complex
2. Formins
What is an Arp in relation to actin microfilaments? What is its function?
Actin-related protein. Arp2 and Arp3 form a complex on which actin monomers can start building a new actin filament.
When Arp2 and Arp3 form a complex to initiate actin nucleation, what end of the new actin filament binds to the complex?
The -ve end (duh).
In what orientation do actin protofilaments twist around each other?
In a right-handed orientation.
How do Arp complexes allow branching nucleation of actin? At what angle from previous actin filaments are new filaments established?
Arp complex binds to existing actin filament and promotes growth of a new filament at a 70 degree angle from the original.
What differentiates the function of an Arp complex from a formin?
Arp: produce branched actin
Formin: produce straight, unbranched actin
How does a formin homodimer interact with an actin filament?
It associates with the growing +ve end and promotes unbranched growth by capturing 2 actin monomers and acting as a template for connecting them to the existing filament.
In vertebrates, what precentage of actin exists as a filament and what percentage is free-floating?
About 50-50%.
Are Arps present in all eukaryotes? Please elaborate.
Yes. Likely arose from an early gene duplication event.
Arps are very similar to actin, so how are they prevented from being co-assembled into a filament?
They have an identical +ve end but a very different -ve end.
What proteins can cap the -ve end of actin?
Arp2, Arp3, tropomodulin.
What proteins can cap the +ve end of actin?
CapZ (plus-end capping protein).
What determines if there is growth or shrinkage of an actin microfilament?
The critical concentration of free actin monomers.
What are the 2 kinds of bundled actin arrangements? What protein is associated with these?
Contractile bundles and tight parallel bundles are associated with formins.
What are the 2 kinds of branched actin arrangements? What protein is associated with these?
Gel-like networks and dendritic networks are associated with Arp complexes.
What protein dimer links actin filaments loosely in a contractile bundle? Why is this advantageous?
α-actinin links actin filaments and allows myosin to enter bundle for muscle contraction.
What protein monomer links actin filaments tightly in a parallel bundle? Why is this advantageous?
Fimbrin links actin filaments and prevents myosin from entering the bundle.
What protein dimer links branched actin filaments in a gel-like network?
Filamin dimers.
What is the structure of a myosin?
They are a coiled-coil of two α-helices with light chains at the N-terminus (neck region) and nothing at the C-terminus.
How many distinct families of myosin exist in humans? How many of these travel towards the negative end of an actin filament?
37 distinct families. Only 1 walks towards the negative end of the filament.
What 5 steps outline the action of myosin 2 with actin?
- Myosin starts bound to actin
- ATP binds, myosin releases
- ATP hydrolyzed to ADP, myosin moves
- Myosin binds actin, releases Pi
- Power stroke, loss of ADP
What phase of myosin movement along actin is associated with rigor mortis?
The point when myosin is bound to actin just before or after a power stroke.
Why is calcium required for muscle contraction?
Calcium binds to the troponin complex, moving tropomyosin off of binding sites and allowing myosin to bind to actin.
How far does myosin II move along an actin filament during each contraction?
~8-9nm.
What causes calcium to be released into the cell during muscle contraction? Where is calcium stored?
An action potential travels down the T-tubules, triggering the release of calcium from the sarcoplasmic reticulum.
Starting at calcium release, outline the steps in the signalling cascade leading to muscle contraction.
- Calcium released from SR
- Calcium binds to calmodulin
- Calmodulin activates myosin light chain kinase
- Phosphorylation of myosin light chain
- Myosins self-assemble into thick filaments for contraction
What differentiates the function of myosin II and myosin V?
Myosin II: muscular contraction
Myosin V: cargo transport along actin filaments
How far does myosin V move along actin during each swing of the lever arm?
~30-40nm.
What are the base monomers of a microtubule?
α-tubulin and β-tubulin.
What is the space in the middle of a microtubule called?
The lumen (same as most biological spaces).
Where does nucleation of a microtubule begin? What protein enables this?
Begins at the microtubule-organizing center (MTOC/centrosome) and is enabled by γ-tubulin small complex (γ-TuSC).
How does γ-tubulin small complex (γ-TuSC) enable microtubule nucleation?
By forming a “lock washer” shape spiral of 7 γ-TuSCs (dimers, 14 γ-tubulins) as a template for polymerization.
How many γ-TuSCs exist on the centrosome matrix?
> 50.
What is a centriole?
A short cylinder of modified microtubules and accessory proteins which exists inside the centrosome matrix of the microtubule organizing center (/centrosome).
Is a centriole required for formation of a centrosome? How can this be tested?
No. By severing a portion of the cell which contains microtubles we can see that they aggregate to form a new centrosome which lacks centrioles.
Actin filaments undergo treadmilling to retain constant length while microtubules have _______ _________.
Dynamic instability.
Describe dynamic instability in microtubles.
Rapid growth with GTP cap followed by loss of GTP cap (catastrophe) and rapid shrinkage. Regaining GTP cap (rescue) allows rapid growth again.
What protein can associate with microtubules and increase the frequency of “catastrophes” or shrinkage events?
Catastrophe factor kinesin-13.
What protein can associate with microtubules and decrease the frequency of “catastrophes” or shrinkage events?
XMAP215.
What effect can MAP2 and tau have on microtubule arrangement when bound to microtubules?
Bind to the side of a microtubule and protrude, causing the microtubules to be more widely spaced.
What are the 2 types of microtubule motor proteins? What direction do they travel along the microtubule?
- Kinesins (travel to +ve end)
2. Dyneins (travel to -ve end)
How does Kinesin move along the microtubules?
It “walks” along. Each step is driven by ATP hydrolysis in the lagging head (“foot”).
What differentiates the structure and function of cytoplasmic dyneins from axonemal dyneins?
Cytoplasmic: homodimers, cargo transport
Axonemal: heterodimers/trimers, cilia and flagella
What drives dynein movement?
ATP hydrolysis.
What is the general structure of a eukaryotic flagella? Do bacteria have the same flagella structure?
9 peripheral microtubule pairs with one central pair. Bacterial flagella have super different structures.
How are dyneins able to drive flagella action and produce movement?
They try to move along the microtubules, but these microtubules are linked so that they only bend a certain amount. Resulting alternation in bending gives whip-like motion.
What are 4 common characteristics of motor proteins?
- Driven by ATP hydrolysis
- Unidirectional movement
- Motor in head
- Tail determines cargo and function
Describe the structure of an intermediate filament.
8 coiled-coil tetramers (32 monomers) are laterally associated and progressively elongated to form a filament.
What are some examples of proteins which can make up intermediate filaments?
Desmin, keratin, neurofilaments, the list goes on…
What part of the cell is most often targeted by toxins? What organism most often employs this strategy?
Plants often produce toxins for defense which target the cytoskeleton.
Do bacteria have actin and microtubules? What about motor proteins?
They have homologs to actin and microtubules but none have motor proteins.
What is the tubulin homolog in bacterial cells?
FtsZ protein.
What are the actin homologs in bacterial cells?
ParM (plasmid segregation), MreB, MbI (cell shape).
What are the 3 main protrusive structures of a cell? What cytoskeletal filament forms these?
All formed by actin:
- Filopodia
- Lamellipodia
- Pseudopodia
Describe a cell’s lamellipodia?
A sheet of actin extending in front of the cell while the back of the cell retracts. A method of cell movement.
Activation of ___ leads to stress fibers, while activation of ___ leads to lamellipodia, and activation of ___ leads to filopodia/microspikes.
Rho = stress fibers, Rac = lamellipodia, Cdc42 = filopodia/microspikes.
How does a neutrophil track down bacteria it wants to engulf?
GPCRs on the membrane detect chemoattractant from the bacterium and trigger actin rearrangement to move the cell.
Which protein associated with actin can bind to actin filaments and to other proteins and activate those upon binding?
Filamin.
