Limits...
A correlative analysis of actin filament assembly, structure, and dynamics.

Steinmetz MO, Goldie KN, Aebi U - J. Cell Biol. (1997)

Bottom Line: Regarding the structure and mechanical properties of the F-actin filament at steady state, no significant correlation with the divalent cation residing in its HAS was found.However, compared to native filaments, phalloidin-stabilized filaments were stiffer and yielded subtle but significant structural changes.Hence, we conclude that the structure and dynamics of the Mg-F-actin moiety within the thin filament are not significantly modulated by the cyclic Ca2+ release as it occurs in muscle contraction to regulate the actomyosin interaction via troponin.

View Article: PubMed Central - PubMed

Affiliation: M.E. Müller Institute for Microscopy, Biozentrum, University of Basel, CH-4056 Basel, Switzerland.

ABSTRACT
The effect of the type of metal ion (i.e., Ca2+, Mg2+, or none) bound to the high-affinity divalent cation binding site (HAS) of actin on filament assembly, structure, and dynamics was investigated in the absence and presence of the mushroom toxin phalloidin. In agreement with earlier reports, we found the polymerization reaction of G-actin into F-actin filaments to be tightly controlled by the type of divalent cation residing in its HAS. Moreover, novel polymerization data are presented indicating that LD, a dimer unproductive by itself, does incorporate into growing F-actin filaments. This observation suggests that during actin filament formation, in addition to the obligatory nucleation- condensation pathway involving UD, a productive filament dimer, a facultative, LD-based pathway is implicated whose abundance strongly depends on the exact polymerization conditions chosen. The "ragged" and "branched" filaments observed during the early stages of assembly represent a hallmark of LD incorporation and might be key to producing an actin meshwork capable of rapidly assembling and disassembling in highly motile cells. Hence, LD incorporation into growing actin filaments might provide an additional level of regulation of actin cytoskeleton dynamics. Regarding the structure and mechanical properties of the F-actin filament at steady state, no significant correlation with the divalent cation residing in its HAS was found. However, compared to native filaments, phalloidin-stabilized filaments were stiffer and yielded subtle but significant structural changes. Together, our data indicate that whereas the G-actin conformation is tightly controlled by the divalent cation in its HAS, the F-actin conformation appears more robust than this variation. Hence, we conclude that the structure and dynamics of the Mg-F-actin moiety within the thin filament are not significantly modulated by the cyclic Ca2+ release as it occurs in muscle contraction to regulate the actomyosin interaction via troponin.

Show MeSH

Related in: MedlinePlus

Schematic representation of F-actin filament polymerization involving the obligatory, UD-based nucleation–condensation pathway and a facultative, LD-based pathway leading to stochastic incorporation of LDs (i.e., via one of their subunits) into  growing F-actin filaments. This polymerization course transiently  yields partially “LD-decorated” filaments. Probably in response  to switching of the filament-bound LDs from a G-like to an F-like  conformation, their unincorporated actin subunits are released  into the monomer pool, thus yielding normal-looking F-actin filaments at steady state.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2141646&req=5

Figure 9: Schematic representation of F-actin filament polymerization involving the obligatory, UD-based nucleation–condensation pathway and a facultative, LD-based pathway leading to stochastic incorporation of LDs (i.e., via one of their subunits) into growing F-actin filaments. This polymerization course transiently yields partially “LD-decorated” filaments. Probably in response to switching of the filament-bound LDs from a G-like to an F-like conformation, their unincorporated actin subunits are released into the monomer pool, thus yielding normal-looking F-actin filaments at steady state.

Mentions: As illustrated schematically in Fig. 9, taken together our polymerization data suggest that during actin filament formation, in addition to the obligatory, UD-based nucleation-condensation pathway, a facultative, LD-based pathway is implicated whose abundance is strongly dependent on the polymerization conditions chosen. Whereas LD-induced branching of polymerizing actin filaments might be a prerequisite for the rapid formation of actin meshworks within distinct compartments (e.g., at the leading edge) of motile cells, stabilization of these filament meshworks is more likely achieved by the concerted action of actin cross-linking proteins. The functional significance of LD is further supported by several recent findings with the actin-binding proteins gelsolin (Hesterkamp et al., 1993) and actobindin (Bubb et al., 1994a,b) and the actin-binding molecule swinholide A (Bubb et al., 1995), all of which appear to stabilize an LD-type actin dimer. In this context, one may also speculate that during dynamic reorganization of the actin cytoskeleton, a substantial amount of actin is present in the form of an LD pool in the cell. However, more detailed studies are now necessary to clarify the exact role of the LD during actin filament assembly and turnover.


A correlative analysis of actin filament assembly, structure, and dynamics.

Steinmetz MO, Goldie KN, Aebi U - J. Cell Biol. (1997)

Schematic representation of F-actin filament polymerization involving the obligatory, UD-based nucleation–condensation pathway and a facultative, LD-based pathway leading to stochastic incorporation of LDs (i.e., via one of their subunits) into  growing F-actin filaments. This polymerization course transiently  yields partially “LD-decorated” filaments. Probably in response  to switching of the filament-bound LDs from a G-like to an F-like  conformation, their unincorporated actin subunits are released  into the monomer pool, thus yielding normal-looking F-actin filaments at steady state.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2141646&req=5

Figure 9: Schematic representation of F-actin filament polymerization involving the obligatory, UD-based nucleation–condensation pathway and a facultative, LD-based pathway leading to stochastic incorporation of LDs (i.e., via one of their subunits) into growing F-actin filaments. This polymerization course transiently yields partially “LD-decorated” filaments. Probably in response to switching of the filament-bound LDs from a G-like to an F-like conformation, their unincorporated actin subunits are released into the monomer pool, thus yielding normal-looking F-actin filaments at steady state.
Mentions: As illustrated schematically in Fig. 9, taken together our polymerization data suggest that during actin filament formation, in addition to the obligatory, UD-based nucleation-condensation pathway, a facultative, LD-based pathway is implicated whose abundance is strongly dependent on the polymerization conditions chosen. Whereas LD-induced branching of polymerizing actin filaments might be a prerequisite for the rapid formation of actin meshworks within distinct compartments (e.g., at the leading edge) of motile cells, stabilization of these filament meshworks is more likely achieved by the concerted action of actin cross-linking proteins. The functional significance of LD is further supported by several recent findings with the actin-binding proteins gelsolin (Hesterkamp et al., 1993) and actobindin (Bubb et al., 1994a,b) and the actin-binding molecule swinholide A (Bubb et al., 1995), all of which appear to stabilize an LD-type actin dimer. In this context, one may also speculate that during dynamic reorganization of the actin cytoskeleton, a substantial amount of actin is present in the form of an LD pool in the cell. However, more detailed studies are now necessary to clarify the exact role of the LD during actin filament assembly and turnover.

Bottom Line: Regarding the structure and mechanical properties of the F-actin filament at steady state, no significant correlation with the divalent cation residing in its HAS was found.However, compared to native filaments, phalloidin-stabilized filaments were stiffer and yielded subtle but significant structural changes.Hence, we conclude that the structure and dynamics of the Mg-F-actin moiety within the thin filament are not significantly modulated by the cyclic Ca2+ release as it occurs in muscle contraction to regulate the actomyosin interaction via troponin.

View Article: PubMed Central - PubMed

Affiliation: M.E. Müller Institute for Microscopy, Biozentrum, University of Basel, CH-4056 Basel, Switzerland.

ABSTRACT
The effect of the type of metal ion (i.e., Ca2+, Mg2+, or none) bound to the high-affinity divalent cation binding site (HAS) of actin on filament assembly, structure, and dynamics was investigated in the absence and presence of the mushroom toxin phalloidin. In agreement with earlier reports, we found the polymerization reaction of G-actin into F-actin filaments to be tightly controlled by the type of divalent cation residing in its HAS. Moreover, novel polymerization data are presented indicating that LD, a dimer unproductive by itself, does incorporate into growing F-actin filaments. This observation suggests that during actin filament formation, in addition to the obligatory nucleation- condensation pathway involving UD, a productive filament dimer, a facultative, LD-based pathway is implicated whose abundance strongly depends on the exact polymerization conditions chosen. The "ragged" and "branched" filaments observed during the early stages of assembly represent a hallmark of LD incorporation and might be key to producing an actin meshwork capable of rapidly assembling and disassembling in highly motile cells. Hence, LD incorporation into growing actin filaments might provide an additional level of regulation of actin cytoskeleton dynamics. Regarding the structure and mechanical properties of the F-actin filament at steady state, no significant correlation with the divalent cation residing in its HAS was found. However, compared to native filaments, phalloidin-stabilized filaments were stiffer and yielded subtle but significant structural changes. Together, our data indicate that whereas the G-actin conformation is tightly controlled by the divalent cation in its HAS, the F-actin conformation appears more robust than this variation. Hence, we conclude that the structure and dynamics of the Mg-F-actin moiety within the thin filament are not significantly modulated by the cyclic Ca2+ release as it occurs in muscle contraction to regulate the actomyosin interaction via troponin.

Show MeSH
Related in: MedlinePlus