Lecture 13

Cytoskeleton: Actin

Actin Filaments

  • Definition: Actin filaments, also known as microfilaments, are helical polymers of the protein actin.

  • Dimensions: Approximately 100 nm in length; actin filaments are about 25 nm in diameter.

Actin Structure

  • Monomers: Each actin subunit is termed G-actin (globular actin), which is a polypeptide that carries either ATP or ADP.

  • Polarity:

    • Plus End (Barbed):

    • ATP-binding pocket is buried within the filament.

    • Minus End (Pointed):

    • ATP-binding pocket is exposed.

  • G-actin vs. F-actin:

    • G-actin refers to individual globular actin monomers.

    • F-actin refers to filamentous actin, which is formed by polymerization of G-actin.

  • Polarity Significance: The assembly of actin monomers gives rise to the filament's polarity, crucial for dynamic processes.

Assembly Dynamics

  • Nucleation: The initial formation (nucleation) of actin filaments is the rate-limiting step. It involves:

    • Weak binding of two actin molecules.

    • Addition of a third actin monomer stabilizes the complex, leading to net elongation at the plus end and loss at the minus end.

  • Treadmilling:

    • Actin treadmilling refers to the dynamic equilibrium where actin filaments lose subunits from the minus end while gaining them at the plus end, effectively pushing the membrane forward in processes like cell migration.

    • This process keeps the filament length relatively stable while the ends are dynamic.

Drugs Affecting Actin Dynamics

  • Phalloidin:

    • A drug derived from the death cap mushroom (Amanita phalloides) that stabilizes F-actin, preventing depolymerization.

    • Can be conjugated with fluorescent markers for visualization.

Actin Structures and Their Functions

  • Cell Structures: Cells dynamically assemble actin filaments into various structures which exhibit different mechanical properties and functionalities:

    • Microvilli

    • Cell cortex

    • Adherens belts

    • Filopodia

    • Lamellipodia (leading edge)

    • Stress fibers

    • Contractile rings

  • Filopodia & Lamellipodia:

    • Filopodia: Protrusions made of tight bundles of actin.

    • Lamellipodia: Broad, branched networks of actin filaments.

Actin-Binding Proteins

  • Role: Actin-binding proteins regulate the polymerization process, length of filaments, and interactions among filaments.

  • Types of Actin-Binding Proteins:

    • Monomer-sequestering proteins (e.g., thymosin ẞ4) prevent G-actin from polymerizing.

    • Actin-polymerizing proteins (e.g., formin) promote the addition of G-actin to filaments.

    • Filament-severing proteins (e.g., gelsolin) cut filaments, thus altering their length.

    • Filament-capping proteins (e.g., CapZ) inhibit G-actin addition and loss at filament ends.

Capping Proteins

  • Properties: Capping proteins are heterodimers resembling two actin monomers that bind to the plus ends of the filaments to regulate polymerization.

  • Capping protein types include CP-α and CP-β.

Actin Crosslinking Proteins

  • Functions:

    • Actin filaments can crosslink to form complex structures such as filopodia and lamellipodia.

    • Alpha-actinin: This is a crosslinking protein making F-actin bundles; it is a rigid dimer with at least two actin-binding sites.

    • Filamin: A flexible crosslinking protein allowing the formation of branched actin networks, facilitating movement and structure adaptability.

  • ERM Proteins (Erzin, Radixin, Moesin): These proteins link the actin cytoskeleton to membranes. They have masked binding sites for actin and membrane proteins that are revealed upon phosphorylation of Erzin.

Branching and Complex Structures

  • Arp2/3 Complex:

    • Composed of 7 subunits resembling G-actin, this complex promotes the branching of actin filaments by binding to existing filaments and recruiting G-actin.

    • Essential for the formation of branched filamentous networks that contribute to cell motility.

Summary of Actin Filament Characteristics

  • G-actin: Globular actin monomers.

  • F-actin: Filamentous actin, characterized by helical structure and ATPase activity.

  • Polarity: Actin filaments are inherently polarized and grow primarily at the plus end, contributing to various cellular functions.

  • Complex Structures: Actin forms various network architectures such as filopodia, lamellipodia, and other structures with differing properties and functions.

  • Actin-Binding Proteins: Functionally diverse, they impact filament growth rates, severing, bundling, and localization to specific cellular locations.