F-Actin assembly

Ikawa T, Hoshino F, Watanabe O, Li Y, Pin-cus P, Safinya CR. (2007) Molecular scale imaging of F-actin assemblies immobilized on a photopolymer surface. Fhys Rev Lett 98, 018101. 

Members of the family of coronin proteins are important for the downregulation of the Arp2/3 complex activity that controls F-actin assembly. Functional work so far has concentrated mainly on coronins 1 (lA), 2 (IB) and 3 (1C) and a significant amount of data has been accumulated yielding first insights into localisation and cellular functions of coronins. 

Tubulin polymerization has several properties in common with the polymerization of actin to form microfilaments. First, at a(3-tubulln concentrations above the critical concentration (CJ, the dimers polymerize into microtubules, whereas at concentrations below the Q, microtubules depolymerlze, similar to the behavior of G-actin and F-actln (see Figure 19-7). Second, the nucleotide, either GTP or GDP, bound to the (3-tubulln causes the critical concentration (CJ for assembly at the (-t) and (—) ends of a microtubule to differ. By analogy with F-actin assembly, the preferred assembly end is designated the (-f) end. Third,... 

Latrunculins A and B are macrolides from the sponge Latrunculia magnified. Latrunculin A (>50 11M) binds close to the nucleotide binding site of G-actin and blocks the assembly with F-actin without promoting disassembly. 

Myosin Subftagment-I Interacts With Two G-Actin Molecules Oligomers of G-Actin and S] Are the Second Intennediates in F-Actin-Si Assembly Conclusion... 

The kinetics of F-actin-Si assembly from G-actin and Si via nucleation of actin filaments, followed by Si binding are not observed in a low ionic strength medium. Instead, the mechanism involves condensation of high affinity (G-actin)2 S complexes rapidly preformed in solution. Assembly of F-actin-Si in the presence of Si > G-actin is a quasi-irreversible process. This mechanism is therefore different from that involving the assembly of F-actin filaments, which is characterized by the initial, energetically unfavorable formation of a small number of nuclei representing a minute fraction of the population of actin molecules, followed by endwise elongation from G-actin subunits. 

Figure 49-3. Schematic representation of the thin fiiament, showing the spatiai configuration of its three major protein components actin, myosin, and tropomyosin. The upper panei shows individual molecules of G-actin. The middle panel shows actin monomers assembled into F-actin. Individual molecules of tropomyosin (two strands wound around one another) and of troponin (made up of its three subunits) are also shown. The lower panel shows the assembled thin filament, consisting of F-actin, tropomyosin, and the three subunits of troponin (TpC, Tpl, andTpT).

Fig. 2.3 The development of polarity and asymmetric division in Saccharomyces cerevisiae. The diagram is reproduced in a slightly simplified form from the work of Lew Reed (1995) with the permission of Current Opinion in Genetics and Development, (a) The F-actin cytoskeleton strands = actin cables ( ) cortical actin patches, (b) The polarity of growth is indicated by the direction of the arrows (arrows in many directions signifies isotropic growth), (c) 10-nm filaments which are assembled to form a ring at the neck between mother and bud. (d) Construction of the cap at the pre-bud site. Notice that the proteins of the cap become dispersed at the apical/isotropic switch, first over the whole surface of the bud, then more widely. Finally, secretion becomes refocussed at the neck in time for cytokinesis, (e) The status and distribution of the nucleus and microtubules of the spindle. Notice how the spindle pole body ( ) plays an important part in orientation of the mitotic spindle.

Bear, J. E., Krause, M. and Gertler, F. B. Regulating cellular actin assembly. Curr. Opin. Cell Biol. 13 158-166, 2001. 

Schmit A-C, Lambert A-M. Microinjected fluorescent phalloidin in vivo reveals the F-actin dynamics and assembly in higher plant mitotic cells. Plant Cell 1990 2 129-138. 

Figure 4.1. Actin polymerisation. Actin monomers (G-actin) may reversibly assemble into actin filaments (F-actin). Profilin binds to G actin (to form profilactin) and thus prevents its polymerisation.

In the resting neutrophil, about 50% of the actin is present in filaments within the cytoskeleton (and hence insoluble in detergents such as Triton X-100), whereas the remainder is detergent soluble and hence is not associated with the cytoskeleton. Data from studies of actin polymerisation in vitro predict that almost all of the actin within the cell should be F-actin (i.e. present in microfilaments). Upon stimulation of neutrophils with agonists such as fMet-Leu-Phe or PMA, actin polymerisation is activated extremely rapidly. There are two important questions Firstly, how is actin maintained in the unpolymerised state in resting cells Secondly, how is it rapidly assembled into the cytoskeleton during activation The answers to these questions lie in understanding the functions of the numerous proteins involved in the assembly and disassembly of actin filaments (Table 4.1). 

Assembly of the actin network merely by interaction with these binding proteins can itself account for pseudopodia formation and propulsive movement. However, there is some evidence to suggest that F-actin-myosin interactions are required for vectorial movement hence it has been demonstrated that pseudopodia contain filament networks comprising actin and myosin. Myosin plays a role in the contractile movement of neutrophils in a... 

The polymer self-assembly theory of Oosawa and Kasai (1962) provides valuable insights into the nature of the nucleation process. The polymerization nucleus is considered to form by the accretion of protomers, but the process is highly cooperative and unfavorable. Indeed, this is strongly suggested by the observation that thousands of actin or tubulin protomers are found in F-actin and microtubule structures if nucleation of self-assembly were readily accomplished and highly favorable, the consequence would be that many more fibers of shorter polymer length would be observed. The Oosawa kinetic theory for nucleation permits one to obtain information about the size of the polymerization nucleus if two basic assumptions can be satisfied in the experimental system. First, the rate of nuclei formation is assumed to be proportional to the loth power of the protomer concentration with io representing the number of protomers required to create the nucleus. Second, the treat-... 

Figure 2 The actin-ADP-ribosylating toxins, (a) Molecular mode of action. The actin-ADP-ribosylating toxins covalently transfer an ADP-ribose moiety from NAD+ onto Arg177 of G-actin in the cytosol of targeted cells. Mono-ADP-ribosylated G-actin acts as a capping protein and inhibits the assembly of nonmodified actin into filaments. Thus, actin polymerization is blocked at the fast-growing ends of actin filaments (plus or barbed ends) but not at the slow growing ends (minus or pointed ends). This effect ultimately increases the critical concentration necessary for actin polymerization and tends to depolymerize F-actin. Finally, all actin within an intoxicated cell becomes trapped as ADP-ribosylated G-actin.

True self-assembly is observed in the formation of many oligomeric proteins. Indeed, Friedman and Beychok reviewed efforts to define the subunit assembly and reconstitution pathways in multisubunit proteins, and all of the several dozen examples cited in their review represent true self-assembly. Polymeric species are also formed by true self-assembly, and the G-actin to F-actin transition is an excellent example. By contrast, there are strong indications that ribosomal RNA species play a central role in specifying the pathway to and the structure of ribosome particles. And it is interesting to note that the assembly of the tobacco mosaic virus (TMV) appears to be a two-step hybrid mechanism the coat protein subunits first combine to form 34-subunit disks by true self-assembly from monomeric and trimeric com-... 

The threshold concentration of monomer that must be exceeded for any observable polymer formation in a self-assembling system. In the context of Oosawa s condensation-equilibrium model for protein polymerization, the cooperativity of nucleation and the intrinsic thermodynamic instability of nuclei contribute to the sudden onset of polymer formation as the monomer concentration reaches and exceeds the critical concentration. Condensation-equilibrium processes that exhibit critical concentration behavior in vitro include F-actin formation from G-actin, microtubule self-assembly from tubulin, and fibril formation from amyloid P protein. Critical concentration behavior will also occur in indefinite isodesmic polymerization reactions that involve a stable template. One example is the elongation of microtubules from centrosomes, basal bodies, or axonemes. 

Figure 5.6 Self-organization in oligomeric proteins. (A) The transacetylase core of the pyruvate dehydrogenase complex. The core consists of 24 identical chains (12 can be seen in this view). (B) The aspartate transcarbamoylase, formed by six catalytic (lighter subunits) and six regulatory chains (darker subunits) (Bi) view showing the threefold symmetry (B2) a perpendicular view. (C) The helical assembly of several identical globular subunits in F-actin polymer. The helix repeats after 13 subunits. (All adapted from Stryer, 1975.)...

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