Capping Protein Increases the Rate of Actin-Based Motility by Promoting Filament Nucleation by the Arp2/3 Complex

Department of Cellular and Molecular Pharmacology, School of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
Cell (Impact Factor: 32.24). 06/2008; 133(5):841-51. DOI: 10.1016/j.cell.2008.04.011
Source: PubMed


Capping protein (CP) is an integral component of Arp2/3-nucleated actin networks that drive amoeboid motility. Increasing the concentration of capping protein, which caps barbed ends of actin filaments and prevents elongation, increases the rate of actin-based motility in vivo and in vitro. We studied the synergy between CP and Arp2/3 using an in vitro actin-based motility system reconstituted from purified proteins. We find that capping protein increases the rate of motility by promoting more frequent filament nucleation by the Arp2/3 complex and not by increasing the rate of filament elongation as previously suggested. One consequence of this coupling between capping and nucleation is that, while the rate of motility depends strongly on the concentration of CP and Arp2/3, the net rate of actin assembly is insensitive to changes in either factor. By reorganizing their architecture, dendritic actin networks harness the same assembly kinetics to drive different rates of motility.

Download full-text


Available from: R. Dyche Mullins, Jan 11, 2014
  • Source
    • "It is a highly conserved protein that is found from yeast to humans, and it is a heterodimer composed of two unrelated subunits, a and b (Wear and Cooper, 2004). Various lines of biochemical evidence (Akin and Mullins, 2008; Loisel et al., 1999) and cellular experiments (Fan et al., 2011; Mejillano et al., 2004; Pappas et al., 2008) have shown that capping protein is essential for various actin-mediated processes. For example, knockdown of capping protein in fast-moving fish keratinocytes causes filopodia-like structures to appear at cell edges (Mejillano et al., 2004), indicating the importance of this protein in cell migration. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Actin polymerization is essential for various stages of mammalian oocyte maturation, including spindle migration, actin cap formation, polar body extrusion, and cytokinesis. The heterodimeric actin-capping protein (CP) is an essential element of the actin cytoskeleton. It binds to the fast-growing (barbed) ends of actin filaments and plays essential roles in various actin-mediated cellular processes. However, the roles of CP in mammalian oocyte maturation are poorly understood. We investigated the roles of CP in mouse oocytes and found that CP is essential for correct asymmetric spindle migration and polar body extrusion. CP mainly localized in the cytoplasm during maturation. By knockdown or ectopically overexpression of CP revealed that CP is critical for efficient spindle migration and maintenance of the cytoplasmic actin mesh density. Expression of the CP inhibiting protein CARMIL impaired spindle migration and polar body extrusion during oocyte maturation and decreased the cytoplasmic actin mesh density. Taken together, these findings show that CP is an essential component of the actin cytoskeleton machinery that plays crucial roles in oocyte maturation, presumably by controlling the cytoplasmic actin mesh density.
    Journal of Cell Science 11/2014; 142(2). DOI:10.1242/jcs.163576 · 5.43 Impact Factor
  • Source
    • "Capping protein (CP) is mandatory for symmetry breaking and motility (Iwasa & Mullins, 2007; Loisel et al., 1999). However, at the concentration used in motility assays (Achard et al., 2010; Akin & Mullins, 2008; Loisel et al., 1999; Reymann et al., 2011), which is usually 20–50 nM, CP activates spontaneous nucleation of actin monomers in solution . Indeed, CP nucleates actin by stabilizing the nuclei at their barbed end leaving them free to elongate from their pointed end (Schafer, Jennings, & Cooper, 1996). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The actin cytoskeleton is a key component of the cellular architecture. However, understanding actin organization and dynamics in vivo is a complex challenge. Reconstitution of actin structures in vitro, in simplified media, allows one to pinpoint the cellular biochemical components and their molecular interactions underlying the architecture and dynamics of the actin network. Previously, little was known about the extent to which geometrical constraints influence the dynamic ultrastructure of these networks. Therefore, in order to study the balance between biochemical and geometrical control of complex actin organization, we used the innovative methodologies of UV and laser patterning to design a wide repertoire of nucleation geometries from which we assembled branched actin networks. Using these methods, we were able to reconstitute complex actin network organizations, closely related to cellular architecture, to precisely direct and control their 3D connections. This methodology mimics the actin networks encountered in cells and can serve in the fabrication of innovative bioinspired systems.
    Methods in enzymology 03/2014; 540:283-300. DOI:10.1016/B978-0-12-397924-7.00016-9 · 2.09 Impact Factor
  • Source
    • "The funneling hypothesis proposes that capping older filaments and inhibiting their growth increases the concentration of available actin monomers, leading to more rapid elongation of new uncapped filaments at the bacterial surface (Carlier et al., 1997). The monomer-gating hypothesis proposes that capping acts as a switch that gates actin monomers to the Arp2/3 complex, enhancing the rate of actin nucleation (Akin and Mullins, 2008). Resolving these hypotheses will await further experimentation. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Intracellular pathogens have developed elaborate mechanisms to exploit the different cellular systems of their unwilling hosts to facilitate their entry, replication, and survival. In particular, a diverse range of bacteria and viruses have evolved unique strategies to harness the power of Arp2/3-mediated actin polymerization to enhance their cell-to-cell spread. In this review, we discuss how studying these pathogens has revolutionized our molecular understanding of Arp2/3-dependent actin assembly and revealed key signaling pathways regulating actin assembly in cells. Future analyses of microbe-host interactions are likely to continue uncovering new mechanisms regulating actin assembly and dynamics, as well as unexpected cellular functions for actin. Further, studies with known and newly emerging pathogens will also undoubtedly continue to enhance our understanding of the role of the actin cytoskeleton during pathogenesis and potentially highlight future therapeutic approaches.
    Cell host & microbe 09/2013; 14(3):242-55. DOI:10.1016/j.chom.2013.08.011 · 12.33 Impact Factor
Show more