Actin Dynamics

 

The fast and reversible polymerization of actin proteins into long filaments consisting of hundreds of subunits is the driving force for many types of cell motions. We are developing models to understand basic properties of actin polymerization in vitro.

 

1. Single actin filament assembly

 

Pure actin filament growth proceeds via polymerization, depolymerization, ATP hydrolysis and release of phosphate. We are interested in understanding how the free energy of ATP hydrolysis is used by cells to power constant turnover of actin filaments ("treadmilling"). In collaboration with I. Fujiwara who measured actin polymerization kinetics in the presence of phosphate, we proposed that the steady state treadmilling of single actin filaments is due to (i) weak affinity of phosphate and ATP for the ends of actin filaments as compared to interior subunits, and (ii) fast phosphate dissociation from terminal subunits.

 

We have also developed a theoretical description of average filament elongation rate and fluctuations, under non-steady state conditions. Length fluctuations were described in terms of the “length diffusivity” which was found to exhibit a pronounced peak below the critical concentration of the barbed end, ccrit (the actin monomer concentration at which net growth vanishes). The peak is due to filaments alternating between capped and uncapped states, a mild version of the dynamic instability of microtubules. This result was compared to observations of large fluctuations above ccrit which questioned the standard picture of monomer-by-monomer elongation (Fujiwara et al. Nature Cell Biol. (2002) 4 666).

     

References

Fujiwara, Vavylonis, and Pollard, Proc. Natl. Acad. Sci. USA 104, 8826–8832 (2007)

Vavylonis, Yang, and O’Shaughnessy, Proc. Natl. Acad. Sci. USA 102, 8543–8548 (2005)

 

2. Formins: regulation of actin polymerization.

 

Formins nucleate actin filaments and remain processively associated at the growing filament end. We developed a quantitative model describing the elongation kinetics of actin filaments associated with formin and profilin. The results were used to interpret parallel experiments by D. R. Kovar et al. (Cell 124, 423 (2006)) who measured actin elongation by TIRF microscopy . The model quantifies how fast formins recruit profilin-actin subunits for polymerization through binding sites in the FH1 domain. The model also rationalizes how filaments associated with formins with many such binding sites can exceed the “diffusion-limited” rate of bare actin.

References

Vavylonis, Kovar, O’Shaughnessy, and Pollard, Molecular Cell, 39 455 (2006).  

 

3. Force generation by polymerization: collective effects in actin bundles

 

Many cellular motions are driven by the polymerization of actin filament bundles growing or shrinking against cellular loads.  We use Brownian ratchet models to study the elongation kinetics of actin bundles polymerizing against an external force.