Intracellular Organization and Movement I: Actin Filaments

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G-actin

Globular actin monomer; ATPase that binds + hydrolyzes ATP; polarity: plus end and minus end.

F-actin

Filamentous actin; double-stranded helix made of G-actin; 7 nm thick; polar.

Actin polarity

Plus end grows ~10x faster than minus end; directionality created by head-to-tail assembly.

ATP–G-actin

Polymerization-favored form; binds to plus end; forms ATP cap during growth.

ADP–G-actin

Depolymerization-favored form; unstable, especially at minus end.

Dynamic instability (actin)

Filament switches between growth + shrinkage based on ATP–G-actin concentration.

Critical concentration (Cc)

Minimum monomer concentration needed for polymerization; different at plus vs minus end.

Treadmilling

Plus end polymerizes while minus end depolymerizes at equal rates → filament length stays constant but “moves.”

Nucleation lag phase

Slow initial formation of actin seed; skipped in cells via nucleating proteins (Arp2/3, formin).

Profilin

Binds ATP–G-actin; promotes polymerization at plus end.

Thymosin β4

Sequesters ATP–G-actin; prevents polymerization.

CapZ

Caps + end of actin; stabilizes; prevents depolymerization.

Tropomodulin

Caps − end; stabilizes; prevents depolymerization.

Tropomyosin

Side-binding protein; stabilizes filament length; blocks myosin binding in muscle.

Cofilin

Severs ADP–F-actin; accelerates depolymerization at minus end; required for lamellipodium rear turnover.

Formin

Nucleates long, unbranched filaments; drives + end elongation (especially filopodia).

Arp2/3 complex

Branches actin at 70°; binds side of preexisting filament; builds lamellipodium network.

Filamin

Cross-links actin at angles → flexible mesh in lamellipodium.

α-actinin

Cross-links parallel actin filaments; found in stress fibers + Z-discs.

Fascin

Bundles actin tightly; essential for filopodia.

Myosin directionality

Generally moves toward the plus end of actin (except myosin VI).

Myosin I

Monomer; vesicle transport + membrane interactions.

Myosin II

Dimer; forms thick filaments in muscle; drives contraction via sliding.

Myosin ATPase cycle – Attach

Myosin bound to actin; no nucleotide.

Myosin ATPase cycle – Release

ATP binds → myosin releases actin.

Myosin ATPase cycle – Cocking

ATP hydrolyzed → ADP + Pi remain bound → hinge bends → head moves ~5 nm.

Myosin ATPase cycle – Power stroke

Binding to actin triggers Pi release → power stroke → ADP leaves.

ATP role in actin-myosin

ATP REQUIRED TO RELEASE myosin from actin, not for power stroke.

Sarcomere

Contractile unit from Z-disc to Z-disc; contains actin + myosin II arrays.

Z-disc

Boundary; α-actinin anchors actin.

Thin filament (actin)

Plus end at Z-disc (CapZ); minus end capped by tropomodulin.

Thick filament (myosin II)

Tail-to-tail dimers; heads point outward.

Nebulin

Stabilizes thin filament length.

Titin

Molecular spring; links myosin to Z-disc; prevents overstretch.

Sliding filament theory

Myosin pulls actin toward midline; filaments do NOT shorten → sarcomere shortens.

SR (sarcoplasmic reticulum)

Stores and releases Ca²⁺ during contraction.

T-tubules

Carry electrical signals deep into the fiber to SR.

Troponin

Binds Ca²⁺ → moves tropomyosin → exposes myosin-binding sites.

Tropomyosin movement

Blocks myosin sites at rest; shifts when Ca²⁺ binds troponin.

Calcium role

Triggers actin–myosin interaction; removed for relaxation.

Smooth muscle regulation

Myosin-regulated, not actin-regulated.

Calmodulin

Ca²⁺ binds to calmodulin.

MLCK

Ca²⁺–calmodulin activates MLCK → phosphorylates myosin light chain → contraction.

Skeletal vs smooth muscle

Skeletal uses troponin/tropomyosin; smooth uses MLCK.

Lamellipodium

Branched actin network at leading edge; pushes membrane forward.

Lamellipodium actin architecture

Arp2/3 branched network + filamin + α-actinin cross-links; capped ends.

Lamellipodium movement

Polymerize at front + cofilin-mediated depolymerization at back.

Filopodia

Thin projections; parallel bundled actin; fascin bundles; formin polymerizes.

Filopodia function

Environmental sensing → guides cell direction.

Focal adhesions

Integrin-based adhesion complexes linking actin to ECM.

Integrins

Bind fibronectin (ECM); connect indirectly to actin.

Fibronectin

ECM glycoprotein bound by integrins.

Adhesion turnover

New adhesions form at front; old ones disassemble at back → required for migration.

Chemotaxis

Directed cell movement based on external signals sensed by filopodia.

Why actin filaments have polarity

Because G-actin monomers are asymmetrical and assemble head-to-tail.

Why polymerization faster at plus end

ATP-G-actin adds preferentially; conformation more favorable.

Actin vs microtubule nucleation

Actin uses Arp2/3 or formin; microtubules nucleate from γ-tubulin ring complex.

Why ATP not used in power stroke

Hydrolysis cocks the head; Pi release generates movement.

Why Ca²⁺ stops contraction when removed

Tropomyosin re-blocks myosin-binding sites.

What cofilin does

Depolymerizes OLD actin (ADP-actin) → essential for migration.

Difference between α-actinin and fascin

α-actinin = loose parallel bundles (lamellipodia/stress fibers); fascin = tight bundles (filopodia).

Explain how Ca²⁺ triggers skeletal muscle contraction.

Ca²⁺ binds troponin → moves tropomyosin → exposes myosin-binding sites → myosin binds actin → power strokes.

Describe the myosin ATPase cycle.

Attach → ATP binds (release) → hydrolysis (cocking) → actin rebinds → Pi release → power stroke → ADP release.

Describe treadmilling.

Plus end polymerizes while minus end depolymerizes at equal rate → filament length constant but monomers move.

Contrast lamellipodia and filopodia.

Lamellipodia = branched (Arp2/3, filamin). Filopodia = bundled (fascin, formin). Different functions.

Explain how fibroblasts migrate.

Lamellipodium polymerizes at front; cofilin depolymerizes at rear; focal adhesions anchor and detach; actin network drives forward movement.

Name all actin-binding proteins involved in lamellipodium formation.

Arp2/3, filamin, α-actinin, CapZ, tropomodulin, cofilin, profilin, thymosin β4.

Describe the structure of the sarcomere.

Z-disc to Z-disc; actin thin filaments anchored at Z-disc; myosin thick filaments in middle; titin connects myosin to Z-disc.

Explain smooth muscle Ca²⁺ regulation.

Ca²⁺ → calmodulin → MLCK → phosphorylates myosin → contraction.

Why don’t actin filaments move inside the lamellipodium?

They are static; growth at front + shrinkage at back creates movement of the network, not individual filaments