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Mathematical modeling of endocytic actin patch kinetics in fission yeast: disassembly requires release of actin filament fragments.

Berro J, Sirotkin V, Pollard TD - Mol. Biol. Cell (2010)

Bottom Line: Conditions inside the cell allow capping protein to bind to the barbed ends of actin filaments and Arp2/3 complex to bind to the sides of filaments faster than the purified proteins in vitro.Simulations also show that depolymerization from pointed ends cannot account for rapid loss of actin filaments from patches in 10 s.An alternative mechanism consistent with the data is that severing produces short fragments that diffuse away from the patch.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06520-8103, USA.

ABSTRACT
We used the dendritic nucleation hypothesis to formulate a mathematical model of the assembly and disassembly of actin filaments at sites of clathrin-mediated endocytosis in fission yeast. We used the wave of active WASp recruitment at the site of the patch formation to drive assembly reactions after activation of Arp2/3 complex. Capping terminated actin filament elongation. Aging of the filaments by ATP hydrolysis and gamma-phosphate dissociation allowed actin filament severing by cofilin. The model could simulate the assembly and disassembly of actin and other actin patch proteins using measured cytoplasmic concentrations of the proteins. However, to account quantitatively for the numbers of proteins measured over time in the accompanying article (Sirotkin et al., 2010, MBoC 21: 2792-2802), two reactions must be faster in cells than in vitro. Conditions inside the cell allow capping protein to bind to the barbed ends of actin filaments and Arp2/3 complex to bind to the sides of filaments faster than the purified proteins in vitro. Simulations also show that depolymerization from pointed ends cannot account for rapid loss of actin filaments from patches in 10 s. An alternative mechanism consistent with the data is that severing produces short fragments that diffuse away from the patch.

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Comparison of experimental and simulated time courses of actin patch assembly and disassembly for the following species: simulated WASp (red curve); measured Wsp1p (red circles); simulated Arp2/3 complex (black curve); measured ARPC5 (black circles); simulated 6% of polymerized actin subunits (teal curve); measured 6% of Act1p (teal triangles); simulated capping protein (green curve); and measured Acp2p (green squares). (A) Simulation of the main model with parameter values measured in biochemical experiments and severing kChop at 0.25 μM−1 s−1. (B) Main model with optimized parameters from Tables 1 and 2. (C) Simulation with optimal parameters from the main model and dissociation of subunits from pointed ends at 0.25 s−1 but without severing and with Arp2/3 complex debranching kDebranch at 0.2 s−1. (D) Same as C but with kDebranch optimized to 0.3 s−1 and actin subunit dissociation rate from pointed ends at 83 s−1 to give the best fit with the experimental data without severing.
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Figure 2: Comparison of experimental and simulated time courses of actin patch assembly and disassembly for the following species: simulated WASp (red curve); measured Wsp1p (red circles); simulated Arp2/3 complex (black curve); measured ARPC5 (black circles); simulated 6% of polymerized actin subunits (teal curve); measured 6% of Act1p (teal triangles); simulated capping protein (green curve); and measured Acp2p (green squares). (A) Simulation of the main model with parameter values measured in biochemical experiments and severing kChop at 0.25 μM−1 s−1. (B) Main model with optimized parameters from Tables 1 and 2. (C) Simulation with optimal parameters from the main model and dissociation of subunits from pointed ends at 0.25 s−1 but without severing and with Arp2/3 complex debranching kDebranch at 0.2 s−1. (D) Same as C but with kDebranch optimized to 0.3 s−1 and actin subunit dissociation rate from pointed ends at 83 s−1 to give the best fit with the experimental data without severing.

Mentions: Because the Wsp1p binding sites and the mechanisms triggering recruitment and activation of Wsp1p are unknown, we simply assumed that recruitment and disappearance of active WASp (reaction 2) follows a Gaussian (see Figure 2D) that drives the other reactions. The simulations give sums of all the WASp molecules in the pathway from reaction 2 to reaction 6 (see red curves, Figures 2 and 3) for comparison with the experimental counts of Wsp1p.


Mathematical modeling of endocytic actin patch kinetics in fission yeast: disassembly requires release of actin filament fragments.

Berro J, Sirotkin V, Pollard TD - Mol. Biol. Cell (2010)

Comparison of experimental and simulated time courses of actin patch assembly and disassembly for the following species: simulated WASp (red curve); measured Wsp1p (red circles); simulated Arp2/3 complex (black curve); measured ARPC5 (black circles); simulated 6% of polymerized actin subunits (teal curve); measured 6% of Act1p (teal triangles); simulated capping protein (green curve); and measured Acp2p (green squares). (A) Simulation of the main model with parameter values measured in biochemical experiments and severing kChop at 0.25 μM−1 s−1. (B) Main model with optimized parameters from Tables 1 and 2. (C) Simulation with optimal parameters from the main model and dissociation of subunits from pointed ends at 0.25 s−1 but without severing and with Arp2/3 complex debranching kDebranch at 0.2 s−1. (D) Same as C but with kDebranch optimized to 0.3 s−1 and actin subunit dissociation rate from pointed ends at 83 s−1 to give the best fit with the experimental data without severing.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2921120&req=5

Figure 2: Comparison of experimental and simulated time courses of actin patch assembly and disassembly for the following species: simulated WASp (red curve); measured Wsp1p (red circles); simulated Arp2/3 complex (black curve); measured ARPC5 (black circles); simulated 6% of polymerized actin subunits (teal curve); measured 6% of Act1p (teal triangles); simulated capping protein (green curve); and measured Acp2p (green squares). (A) Simulation of the main model with parameter values measured in biochemical experiments and severing kChop at 0.25 μM−1 s−1. (B) Main model with optimized parameters from Tables 1 and 2. (C) Simulation with optimal parameters from the main model and dissociation of subunits from pointed ends at 0.25 s−1 but without severing and with Arp2/3 complex debranching kDebranch at 0.2 s−1. (D) Same as C but with kDebranch optimized to 0.3 s−1 and actin subunit dissociation rate from pointed ends at 83 s−1 to give the best fit with the experimental data without severing.
Mentions: Because the Wsp1p binding sites and the mechanisms triggering recruitment and activation of Wsp1p are unknown, we simply assumed that recruitment and disappearance of active WASp (reaction 2) follows a Gaussian (see Figure 2D) that drives the other reactions. The simulations give sums of all the WASp molecules in the pathway from reaction 2 to reaction 6 (see red curves, Figures 2 and 3) for comparison with the experimental counts of Wsp1p.

Bottom Line: Conditions inside the cell allow capping protein to bind to the barbed ends of actin filaments and Arp2/3 complex to bind to the sides of filaments faster than the purified proteins in vitro.Simulations also show that depolymerization from pointed ends cannot account for rapid loss of actin filaments from patches in 10 s.An alternative mechanism consistent with the data is that severing produces short fragments that diffuse away from the patch.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06520-8103, USA.

ABSTRACT
We used the dendritic nucleation hypothesis to formulate a mathematical model of the assembly and disassembly of actin filaments at sites of clathrin-mediated endocytosis in fission yeast. We used the wave of active WASp recruitment at the site of the patch formation to drive assembly reactions after activation of Arp2/3 complex. Capping terminated actin filament elongation. Aging of the filaments by ATP hydrolysis and gamma-phosphate dissociation allowed actin filament severing by cofilin. The model could simulate the assembly and disassembly of actin and other actin patch proteins using measured cytoplasmic concentrations of the proteins. However, to account quantitatively for the numbers of proteins measured over time in the accompanying article (Sirotkin et al., 2010, MBoC 21: 2792-2802), two reactions must be faster in cells than in vitro. Conditions inside the cell allow capping protein to bind to the barbed ends of actin filaments and Arp2/3 complex to bind to the sides of filaments faster than the purified proteins in vitro. Simulations also show that depolymerization from pointed ends cannot account for rapid loss of actin filaments from patches in 10 s. An alternative mechanism consistent with the data is that severing produces short fragments that diffuse away from the patch.

Show MeSH
Related in: MedlinePlus