#actin polymerization

Regeneration -ll

Actin polymerization in sperm cells occurs primarily during:

Actin polymerization in sperm is temporally restricted to acrosomal reaction. G-actin monomers stored bound to profilin in sperm head rapidly assemble into F-actin bundle forming slender acrosomal process that extends bindin to reach vitelline envelope. Calcium influx and pH rise activate actin nucleation via formin and depolymerization of capping proteins, generating force to protrude process through egg jelly remnants. Cytochalasin B or latrunculin blocking polymerization prevents process formation and fertilization fails. Cortical reaction involves secretory exocytosis in egg, fertilization cone actin polymerization occurs in egg cortex during sperm incorporation, and slow block relies on envelope hardening rather than actin assembly.

Ref: Gilbert, Developmental Biology, 12th ed., Chapter 7: Actin polymerization driving acrosomal process extension in sea urchin sperm.

View Details

Sea urchin -l

Actin polymerization in sperm cells occurs primarily during:

During acrosomal reaction, sperm subacrosomal region contains concentrated pool of globular actin monomers maintained unpolymerized by profilin. Upon Ca2+ influx triggered by egg jelly contact, actin rapidly polymerizes into bundle of filamentous F-actin pushing acrosomal membrane forward forming slender elongated acrosomal process about one micrometer long, thrusting bindin-bearing tip through viscous egg jelly toward vitelline envelope receptor field. Cytochalasin D experiments blocking actin polymerization abolish process extension and prevent fertilization. Fertilization cone actin polymerization occurs in egg cortex, cortical reaction involves egg exocytosis, not sperm actin. Thus actin polymerization in sperm cells occurs primarily during acrosomal reaction creating penetration organelle.

Ref: Tilney & Inoue, J Cell Biol 1985, Acrosomal process actin; Gilbert Chapter 7: Actin polymerization in sperm.

View Details

Cytoskeleton

Which protein is responsible for the intracellular movement of Listeria by actin polymerization?

Intracellular pathogenesis of Listeria monocytogenes illustrates actin based propulsion mechanism. After internalization via E cadherin or Met receptor mediated endocytosis, bacterium escapes vacuole via pore forming toxin listeriolysin O and phospholipases PlcA PlcB, enters cytosol and replicates. It then expresses ActA protein anchored via C terminal transmembrane domain, retained at old pole after secretion. ActA N terminal domain mimics eukaryotic nucleation promoting factor recruiting and activating host Arp2/3 complex via acidic motifs and binding Ena VASP via proline rich repeats accelerating elongation. Activation produces dense branched network forming actin tail up to 10 micron long comet behind bacterium with barbed ends oriented toward bacterial surface. Continuous polymerization generates compressive force propelling bacterium forward at approximately 0.1 to 1 micron per second, enabling protrusion into neighboring cell forming double membrane secondary vacuole. Isogenic actA deletion mutants avirulent immobile in cytosol. Capping protein regulates tail length but not initiation, tropomyosin stabilizes stress fibers, vinculin adhesion linking, none responsible for Listeria propulsion mechanism.

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 16: Listeria ActA and Actin-Based Motility.

View Details

Cytoskeleton

The polymerization of actin filaments follows which sequence?

Actin assembly exhibits sigmoidal kinetics reflecting cooperative nucleation mechanism. Lag phase corresponds to nucleation where monomers collide forming unstable dimer with Kd micromolar and trimer tetramer nucleus, high free energy barrier due to entropy loss and weak contacts, hence extremely slow without nucleators. Over minutes trimer accumulates reaching critical nucleus size three to four subunits, providing template for rapid elongation phase where ATP G actin adds efficiently at barbed end with near diffusion limited rate, elongating filament linearly until monomer pool depleted to critical concentration. Eventually steady state reached where on rate equals off rate at both ends, total polymer mass constant while individual subunits treadmill hydrolyzing ATP, known as steady state treadmilling. This order nucleation elongation steady state parallels crystallization, microtubule assembly and amyloid formation. Alternative orders reversing steps violate thermodynamic principle that stable nucleus must form before rapid growth. Pharmacological inhibitors target stages: latrunculin sequesters monomer preventing nucleation, cytochalasin D caps barbed end blocking elongation.

Ref: Lodish et al., Molecular Cell Biology, 9th ed., Chapter 18: Actin Assembly Nuceleation Elongation Stages.

View Details

Cytoskeleton

The rate of ATP-actin addition is __ times faster at the (+) end compared to the (-) end.

Elongation kinetics at filament ends measured using TIRF microscopy of single filaments and pyrene actin assembly shows strong asymmetry. Barbed plus end characterized by high on rate constant 11.6 per micromolar per second and off rate 1.4 per second, critical concentration 0.12 micromolar, while pointed minus end on rate 1.3 per micromolar per second off rate 0.8 per second Cc 0.6 micromolar. Ratio on rates roughly nine fold, often approximated as ten fold faster at plus end for ATP actin at physiological ionic strength 50 millimolar KCl 1 millimolar MgCl2. ATP actin cap at plus end stabilizes, with ATP hydrolysis occurring after incorporation generating ADP Pi intermediate that still retains higher affinity than ADP. ADP actin at minus end less stable. Profilin ATP actin addition exclusive to plus end exaggerates bias in vivo. This kinetic difference drives treadmilling when free monomer between two Cc values, enables lamellipodial protrusion where plus ends face membrane, and underlies polarity dependent myosin directionality vital for cell migration and contractility.

Ref: Pollard and Earnshaw, Cell Biology, Chapter 12: Barbed End Kinetics and Actin Polarity.

View Details