Cell Biology (288) -- Unit 7: Cytoskeletal Systems

Front 1

3 main components of cytoskeleton:

Back 1

1) microtubules (25 nm diameter)
2) microfilaments (7 nm diameter)
3) intermediate filaments (10 nm diameter)

Front 2

Where do microtubules come from?

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Emanate from the centrosome of the nucleus.

Front 3

How are microfilaments organized in the cell?

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Often run directionally through the cytosol and circularly around the perimeter of the cell.

Front 4

Where to intermediate filaments come from and how are they organized?

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Emanate centrally but with no organizational pattern.

Front 5

What do microtubules do? (3)

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1) Give structural organization to the cell
2) Form the spindle poles of the mitotic spindle
3) Grow from basal bodies into the cilia of cells

Front 6

What is the monomeric unit of microtubules?

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Tubulin: alpha and beta tubulin heterodimers that are structurally almost identical but only 40% identical at the AA level.

Front 7

What is the structure of a tubulin monomer?

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Each monomer (two subunits) has a GTP-binding domain at the N-terminal, a central domain where microtubule toxins bind, and a C-terminal domain where microtubule-associated proteins interact.

Front 8

How do microtubules assemble?

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1) Basal body is used as a nucleus.
2) Monomers assemble in a linear array to form a protofilament-- beta end is + (faster) and alpha end is - (slower).
3) 13 protofilaments form a hollow microtubule.

Front 9

What does microtubule assembly require?

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-tubulin
-GTP (allosteric [conformation] regulator, not energy source)
-basal body
-magnesium

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Elongation phase:

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High concentration of GTP-bound tubulin causes rapid assembly from both + and - ends.

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Plateau phase:

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Treadmilling: tubulin added = tubulin lost. Dependent on critical concentration of tubulin, which is lower at + than at - end (so tubulin is added at + when it is being lost at -).

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Dynamic instability model:

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rapid mictrotubule growth followed by collapse (or catastrophe). Enabled by the fact that GTP, which makes the tubulin monomers more stable, eventually degrades to GDP, which is not stabilizing.

Front 13

Microtubules in red blood cells:

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RBC eject their nuclei and centrosomes, causing microtubules to form a peripheral ring of cortical bundles for structure.

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Centrosome:

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microtubule-organizing centre (MTOC). Two perpendicular centrioles surrounded by a sheath that has pores (nucleating sites) formed by rings of gamma-tubulin from which microtubules emerge.

Front 15

Microfilaments:

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Also actin filaments, F-actin.

Front 16

What do microfilaments do? (3)

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1) provide cell structure
2) play a role in amoeboid movement (by actin polymerization and contraction)
3) provide structural support in microvilli

Front 17

What is the monomeric unit of microfilaments?

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Monomer is G-actin (globular actin), a U-shaped protein with an ATP binding site at the centre. There are beta, alpha and gamma actin isoforms in the cell-- beta most common.

Front 18

Microfilament assembly:

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A lot like microtubule assembly:
-ATP as an allosteric regulator (instead of GTP)
-has a + and - end dependent on different critical concentrations
-treadmilling
-forms a twisted coil of two filaments

-bundling proteins link microfilaments into bundles
-also proteins for capping (to end polymerization and prevent catastrophe), nucleating, side-binding, sequestering G-actin (to prevent polymerization)

Front 19

Intermediate filaments:

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Interconnected to other cytoskeletal elements and ECM with protein crossbridges.

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What is the monomeric unit of intermediate filaments?

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Monomeric unit is alpha helical monomer with globular domains at N- and C-terminals.

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How does intermediate filament assembly work? (5)

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1) Monomers become coiled-coil dimers that retain polarity of the monomer.
2) Two dimers form a staggered anti-parallel tetramer that lacks polarity.
3) Tetramer is the monomer of the filament strand.
4) Many filament strands can be twisted together to form a filament.
5) Once created, filaments exist until the cell dies.

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What role do intermediate filaments often play in the cell?

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Creating flexible impermeable water barriers.

Front 23

Cellular motility:

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Movements that occur at the tissue level via skeletal muscle are actually large-scale coordinated cellular movements, which are based on sub-cellular movements, which are based on molecular movements of motor proteins and mechano enzymes.

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Two types of cytosolic subcellular transport:

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1) Anterograde: with kinesins, from the cell body to the synapse or membrane (to the PLUS end) (dominant over retrograde).
2) Retrograde: with dyneins, from the synapse or membrane to the cell body (to the MINUS end).

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Kinesin:

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Coiled alpha-helix stalk with two globular heads driven by ATP hydrolysis, and with a tail full of light chains that interact with the cargo.

Globular heads interact with the beta-tubulin subunits in the microtubule.

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Dynein:

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Two heavy chains with globular head whose stalks interact with the microtubule. Heavy chains attach to stems, which have light chains attached to the cargo.

Front 27

Two microtubule-based motors for axonemic subcellular transport:

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1) Cilia: short, many; move in a whiplike motion.
2) Flagella: long, singular; move in a wavelike motion.

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Basal body:

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A pinwheel of microtubule triplets, one complete and two partial, connected to each other as well as two the central region via radial spokes.

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Axoneme:

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A cilium or flagellum. Structurally, they are the same: microtubules growing out of the basal body in a 9+2 pattern that slide past each other to allow the axoneme to flex.

Inner and outer dynein arms hydrolyze ATP and move the axoneme by contracting, pulling on the beta tubule of the neighbouring microtubule doublet.

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Two categories of motility:

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1) Muscle
2) Non-muscle

Front 31

Muscle motility:

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Needs type II myosin, which has 2 heavy chains, each with a large globular head and ATP catalytic site, coiled around each other.

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Non-muscle motility:

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-intracellular vesicular transport, cytoplasmic streaming, amoeboid movement, cell growth, cell shape

Uses type V myosin, which is very much like type II but has more light chains behind the globular heads, and has a tail that interacts with intermediate proteins (e.g. RABs-- cell surface and vesicle markers) that hold the cargo vesicle.

Type V myosin walks on microfilaments like kinesin on microtubules. Microfilaments are twisted, with 13 subunits per turn (36 nm long)-- so myosin V takes long steps.

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Axonemic cellular movement:

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Extension, contact, grasping, pulling.

Done with lamellipodia and filopodia (thin cytoplasmic streams ahead of the leading edge of the lamellipodium).