Detailed Cytoskeleton Notes

Cytoskeleton

Overview

• The cytoskeleton is a network of protein filaments within cells.
• It provides structural support, facilitates transport, and plays a role in cell division.
• All cells possess a cytoskeleton, but this discussion focuses on animal cells due to their lack of a cell wall, making the cytoskeleton’s role more critical.

Major Types of Cytoskeletal Proteins

• Three major types:
  • Intermediate Filaments: Provide mechanical strength and stability.
  • Microtubules: Involved in intracellular transport, cell shape and support, movement (e.g., flagella), and chromosome segregation during cell division.
  • Actin Filaments (Microfilaments): Contribute to cell shape, motility, contraction, and cell division.

Intermediate Filaments

• The strongest cytoskeletal filaments.
• Their name reflects their intermediate diameter.
• Protect cells from mechanical and torsional stress.
• Located in the cytoplasm of animal cells.
• Contribute to cell-cell junctions (desmosomes).
• Form the nuclear lamina, reinforcing the nuclear envelope.

Structure

• Monomers with alpha-helical secondary structures.
• Alpha-helices form dimeric coiled-coils.
• Two dimeric coiled-coils create staggered antiparallel tetramers.
  • These tetramers run in opposite directions.
  • Possess an unstructured N-terminal overhang.
• Interactions are noncovalent, except for the primary sequence.
• Tetramers associate laterally via noncovalent interactions, forming bundles of 8 tetramers (32 monomers).
• Bundles interact through protein overhangs, allowing growth in both directions.
• Intermediate filaments lack polarity (both ends are the same).
• Rope-like structure contributes to strength and resistance to mechanical stress.

Classes

• Different cells/locations have different intermediate filaments.
• Amino acid sequence of coiled-coil domains is conserved, but overhangs vary, leading to different types.
• Cytoplasmic intermediate filaments show diversity based on cell type.
• Nuclear lamins are unique to the nucleus.

Keratins

• Found in all epithelial cells.
• Over 50 types exist, each with unique roles.
• Can be anchored in desmosomes (cell-cell junctions).
• Can be secreted to form structures like nails.

Neurofilaments

• Present in neuron axons.
• Provide stability to axons.
• Increased neurofilament levels correlate with ALS development, potentially contributing to neuronal damage.

Nuclear Lamina

• Intermediate filaments in the nucleus are called lamins.
• Form a meshwork to reinforce the nuclear envelope.
• Broken down by kinases (phosphorylation) during cell division and reformed by protein phosphatases after cell division.
• Nuclear lamina problems are linked to diseases like progeria (premature aging).
• Potential link to normal aging and altered chromosome positioning is investigated.

Accessory Proteins: Plectin

• Bundle or connect intermediate filaments to other cytoskeletal proteins.
• Plectin binds intermediate filaments to microtubules.
• Facilitates interactions between intermediate filaments and other proteins.

Linker Proteins

• Link cytoskeletal proteins to each other.
• SUN-domain proteins bind to the nuclear lamina and interact with KASH-domain proteins.
• This interaction creates an indirect connection between nuclear and cytosolic cytoskeletal proteins.

Microtubules

• Help organize eukaryotic cells.
• Located throughout the cytoplasm.
• Serve as tracks for transporting vesicles, organelles, and macromolecules.
• Dynamic structures that rapidly assemble and disassemble based on cellular needs.
• Also form stable structures like cilia and flagella.

Structure

• Built from α and β tubulin proteins.

• α and β tubulin interact non-covalently to form tubulin dimers (microtubule subunits).

• Subunits stack non-covalently to form protofilaments.

• Microtubules have polarity: a plus end (β-tubulin) and a minus end (α-tubulin).

• 13 protofilaments form a tube-like structure.

• Noncovalent interactions between protofilaments.

Microtubule Organizing Centers (MTOCs)

• Tubulin dimers polymerize into microtubules with the help of MTOCs.
• γ-tubulin ring complex (containing 13 tubulin dimers) forms at the MTOC to initiate microtubule formation.

Centrosome

• Common MTOC in animal cells.
• Consists of two centrioles surrounded by a gel-like protein matrix.
• Located near the nucleus.
• The plus end of the microtubule extends into the cytoplasm, while the minus end faces the centrosome matrix.
• Microtubule nucleation begins at the γ-tubulin ring.

Other Examples

• γ-tubulin rings can form at other locations besides the centrosome, depending on cell type (e.g., nuclear envelope in yeast, apical side of intestinal epithelial cells).

Dynamic Structures

• Microtubules grow rapidly from the organizing site, then shrink or disappear.
• Growth is independent of other microtubules, known as dynamic instability.
• Dynamic instability allows microtubules to explore the cell.

Cause of Dynamic Instability

• Tubulin dimers bind GTP.
• Tubulin dimers hydrolyze GTP to GDP.
• Rapid growth occurs when growth is faster than hydrolysis.
• Tubulin dimers bound to GTP form a GTP cap at the end of the microtubule, creating a stable structure.
• As growth slows, hydrolysis outpaces growth.
• Tubulin-GDP is less stable, causing the microtubule to disassemble.

Regulation by Microtubule Associated Proteins (MAPs)

• Microtubules are unstable during cell division, providing tubulin dimers for the mitotic spindle.
• MAPs stabilize microtubules or link them to other cytoskeletal components.
• The γ-tubulin ring complex helps nucleate microtubules.
• Some proteins stabilize microtubules, while others (like augmin) induce branching.
• Other proteins promote polymerization.

As a Cellular Highway

• Random diffusion is too slow for efficient intracellular transport.
• Microtubules act as tracks for movement of materials.
• Movement can occur toward or away from the nerve cell body.

Movement Along Microtubules

• Motor proteins move materials along microtubules.
• Two major classes:
  • Kinesins: Move toward the plus end.
  • Dyneins: Move toward the minus end.
• Both have globular heads (associated with microtubules) and tails (interact with cargo).
• The tail's amino acid sequence determines cargo specificity.

Motor Proteins Move By Hydrolyzing ATP

• Globular heads bind ATP.
• ATP binding increases affinity for the microtubule.
• ATP hydrolysis causes them to loosen up and move toward the positive or negative pole, depending on the motor protein type (kinesin or dynein).
• The motor proteins effectively “walk” along the microtubule.

Transport Mechanism

• Cargo is attached directly to the tails or via adaptor proteins.
• Microtubules are busy highways, with kinesins and dyneins moving in opposite directions simultaneously.

Cilia

• Eukaryotic cells have cilia on their surface (e.g., protists, respiratory tract cells).
• Cilia have stable microtubules originating from a basal body.
• Move in a whip-like fashion (power stroke, recovery step).

Flagella

• Enable cell movement in aqueous environments.
• Longer than cilia and move the whole cell, not just the fluid.
• Move in rhythmic motions.
• Bacterial flagella function differently (convergent evolution).

How Cilia and Flagella Move

• 9 + 2 arrangement of microtubules (9 microtubule doublets around the outside, two in the center).
• Microtubules are connected by linking proteins.
• Movement is driven by a specialized dynein.

Movement of Dynein in Cilia and Flagella

• Dynein on one of the two microtubules causes sliding via ATP hydrolysis.
• Linking proteins cause bending rather than sliding.
• Bending causes cilia movement or flagella-driven swimming.
• All 9 pairs move in the same way at once.

Actin or Microfilaments

• Made up of the protein actin.
• Form structures like villi in the intestine.
• Help maintain cell shape.
• Aid in cell division (cytokinesis).
• Help in cell motility.
• Frequently associated with microtubules.

Structure

• Actin is a globular protein (5% of total protein mass).
• Half is free, and half is in filaments.
• Has polarity like microtubules.
• Thinner and shorter than microtubules, but more numerous.
• Flexible.

Assembly

• Similar assembly to microtubules.
• Actin monomer binds ATP (instead of GTP).
• ATP-bound monomers add to both ends (plus end addition is faster).
• ATP is hydrolyzed after binding, destabilizing the filament and releasing the monomer.

Treadmilling

• Occurs when actin monomers are not in excess.
• Addition at the plus end is faster than ATP hydrolysis.
• Hydrolysis is faster at the minus end than addition.
• Filament length remains constant, but monomers “move through.”

Actin Binding Proteins

• Control actin function.
• Can sequester monomers to prevent filament formation.
• Can nucleate filament formation (ARP complex).
• Crosslink filaments, cap filaments, and function as motor proteins.

Myosins

• Motor proteins that move along actin fibers.
• Use ATP hydrolysis for movement.
• Myosin I and Myosin II are common subfamilies found in animal cells.

Myosin I Movement

• Head domains hydrolyze ATP and detach until rebinding ATP.
• Move along actin fibers toward the plus pole.
• Cargo is determined by tail amino acids.
• Can move vesicles or reshape the plasma membrane.

Cell Cortex

• Located beneath the plasma membrane, providing structure and strength.
• Loose meshwork in RBCs for deformability, tighter in most other cells.
• Actin fibers are anchored to the plasma membrane by attachment proteins.

Actin Rearrangement In Cell Movement

• Some cells (immune cells, unicellular eukaryotes) move by actin rearrangements, particularly when attracted to chemical signals.

• Three main steps:

  1. Cell sends out protrusions at the “front” or leading edge.
  2. Protrusions adhere to the surface.
  3. The rest of the cell drags itself forward by traction on these points of anchorage.