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Mechanism of Cdc42-induced actin polymerization in neutrophil extracts.

Zigmond SH, Joyce M, Yang C, Brown K, Huang M, Pring M - J. Cell Biol. (1998)

Bottom Line: Electron microscopy revealed that Cdc42-induced filaments elongated rapidly, achieving a mean length greater than 1 micron in 15 s.There was little change in mean length of Cdc42-induced filaments between 15 s and 5 min, suggesting that the increase in F-actin over this time was due to an increase in filament number.These data suggest that Cdc42 induction of actin polymerization requires both creation of free barbed ends and facilitated elongation at these ends.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Pennsylvania. Philadelphia, Pennsylvania 19104-6018, USA. szigmond@sas.upenn.edu

ABSTRACT
Cdc42, activated with GTPgammaS, induces actin polymerization in supernatants of lysed neutrophils. This polymerization, like that induced by agonists, requires elongation at filament barbed ends. To determine if creation of free barbed ends was sufficient to induce actin polymerization, free barbed ends in the form of spectrin-actin seeds or sheared F-actin filaments were added to cell supernatants. Neither induced polymerization. Furthermore, the presence of spectrin-actin seeds did not increase the rate of Cdc42-induced polymerization, suggesting that the presence of Cdc42 did not facilitate polymerization from spectrin-actin seeds such as might have been the case if Cdc42 inhibited capping or released G-actin from a sequestered pool. Electron microscopy revealed that Cdc42-induced filaments elongated rapidly, achieving a mean length greater than 1 micron in 15 s. The mean length of filaments formed from spectrin-actin seeds was <0.4 micron. Had spectrin-actin seeds elongated at comparable rates before they were capped, they would have induced longer filaments. There was little change in mean length of Cdc42-induced filaments between 15 s and 5 min, suggesting that the increase in F-actin over this time was due to an increase in filament number. These data suggest that Cdc42 induction of actin polymerization requires both creation of free barbed ends and facilitated elongation at these ends.

Show MeSH
A Model. Cdc42 is depicted as interacting with several  factors including Nuc that creates free barbed ends and an elongation promoting factor (EP) that facilitates rapid elongation.  Filament elongation is terminated by release of EP and/or addition of a capper.
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Figure 9: A Model. Cdc42 is depicted as interacting with several factors including Nuc that creates free barbed ends and an elongation promoting factor (EP) that facilitates rapid elongation. Filament elongation is terminated by release of EP and/or addition of a capper.

Mentions: Exactly how Cdc42 promotes polymerization is not clear. However, a model consistent with our data is shown in Fig. 9: Cdc42 activates a complex which both creates a free barbed end (Nuc) and promotes elongation (EP) of that end. We suggest that Cdc42 remains with the elongation-promoting complex, thus acting in manner parallel to ActA in Listeria movement. Alternatively, Cdc42 could merely activate the elongation-promoting complex. The promotion of elongation involves enhanced rates of G-actin addition (we cannot rule out the possibility that it also allows prolonged elongation). After a short time the rapid elongation is terminated either by release of the elongation promoting complex and/or by capping. The activated Cdc42 can then assemble a new active complex.


Mechanism of Cdc42-induced actin polymerization in neutrophil extracts.

Zigmond SH, Joyce M, Yang C, Brown K, Huang M, Pring M - J. Cell Biol. (1998)

A Model. Cdc42 is depicted as interacting with several  factors including Nuc that creates free barbed ends and an elongation promoting factor (EP) that facilitates rapid elongation.  Filament elongation is terminated by release of EP and/or addition of a capper.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132886&req=5

Figure 9: A Model. Cdc42 is depicted as interacting with several factors including Nuc that creates free barbed ends and an elongation promoting factor (EP) that facilitates rapid elongation. Filament elongation is terminated by release of EP and/or addition of a capper.
Mentions: Exactly how Cdc42 promotes polymerization is not clear. However, a model consistent with our data is shown in Fig. 9: Cdc42 activates a complex which both creates a free barbed end (Nuc) and promotes elongation (EP) of that end. We suggest that Cdc42 remains with the elongation-promoting complex, thus acting in manner parallel to ActA in Listeria movement. Alternatively, Cdc42 could merely activate the elongation-promoting complex. The promotion of elongation involves enhanced rates of G-actin addition (we cannot rule out the possibility that it also allows prolonged elongation). After a short time the rapid elongation is terminated either by release of the elongation promoting complex and/or by capping. The activated Cdc42 can then assemble a new active complex.

Bottom Line: Electron microscopy revealed that Cdc42-induced filaments elongated rapidly, achieving a mean length greater than 1 micron in 15 s.There was little change in mean length of Cdc42-induced filaments between 15 s and 5 min, suggesting that the increase in F-actin over this time was due to an increase in filament number.These data suggest that Cdc42 induction of actin polymerization requires both creation of free barbed ends and facilitated elongation at these ends.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Pennsylvania. Philadelphia, Pennsylvania 19104-6018, USA. szigmond@sas.upenn.edu

ABSTRACT
Cdc42, activated with GTPgammaS, induces actin polymerization in supernatants of lysed neutrophils. This polymerization, like that induced by agonists, requires elongation at filament barbed ends. To determine if creation of free barbed ends was sufficient to induce actin polymerization, free barbed ends in the form of spectrin-actin seeds or sheared F-actin filaments were added to cell supernatants. Neither induced polymerization. Furthermore, the presence of spectrin-actin seeds did not increase the rate of Cdc42-induced polymerization, suggesting that the presence of Cdc42 did not facilitate polymerization from spectrin-actin seeds such as might have been the case if Cdc42 inhibited capping or released G-actin from a sequestered pool. Electron microscopy revealed that Cdc42-induced filaments elongated rapidly, achieving a mean length greater than 1 micron in 15 s. The mean length of filaments formed from spectrin-actin seeds was <0.4 micron. Had spectrin-actin seeds elongated at comparable rates before they were capped, they would have induced longer filaments. There was little change in mean length of Cdc42-induced filaments between 15 s and 5 min, suggesting that the increase in F-actin over this time was due to an increase in filament number. These data suggest that Cdc42 induction of actin polymerization requires both creation of free barbed ends and facilitated elongation at these ends.

Show MeSH