scholarly journals Inhibition of polar actin assembly by astral microtubules is required for cytokinesis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Anan Chen ◽  
Luisa Ulloa Severino ◽  
Thomas C. Panagiotou ◽  
Trevor F. Moraes ◽  
Darren A. Yuen ◽  
...  
Keyword(s):  
Cell Division ◽  
Actin Cytoskeleton ◽  
Actin Filament ◽  
Cell Cortex ◽  
Actin Assembly ◽  
Actin Network ◽  
Contractile Ring ◽  
Actin Networks ◽  
Membrane Blebbing ◽  
Actin Isoform

AbstractDuring cytokinesis, the actin cytoskeleton is partitioned into two spatially distinct actin isoform specific networks: a β-actin network that generates the equatorial contractile ring, and a γ-actin network that localizes to the cell cortex. Here we demonstrate that the opposing regulation of the β- and γ-actin networks is required for successful cytokinesis. While activation of the formin DIAPH3 at the cytokinetic furrow underlies β-actin filament production, we show that the γ-actin network is specifically depleted at the cell poles through the localized deactivation of the formin DIAPH1. During anaphase, CLIP170 is delivered by astral microtubules and displaces IQGAP1 from DIAPH1, leading to formin autoinhibition, a decrease in cortical stiffness and localized membrane blebbing. The contemporaneous production of a β-actin contractile ring at the cell equator and loss of γ-actin from the poles is required to generate a stable cytokinetic furrow and for the completion of cell division.

scholarly journals MICAL2 acts through Arp3B isoform-specific Arp2/3 complexes to destabilize branched actin networks

2020 ◽  
Author(s):  
Chiara Galloni ◽  
Davide Carra ◽  
Jasmine V. G. Abella ◽  
Svend Kjær ◽  
Pavithra Singaravelu ◽  
...  
Keyword(s):  
Functional Properties ◽  
Actin Filament ◽  
Actin Assembly ◽  
Actin Network ◽  
Network Stability ◽  
Actin Networks ◽  
Cellular Processes ◽  
Actin Turnover ◽  
Filament Networks

AbstractThe Arp2/3 complex (Arp2, Arp3 and ARPC1-5) is essential to generate branched actin filament networks for many cellular processes. Human Arp3, ARPC1 and ARPC5 exist as two isoforms but the functional properties of Arp2/3 iso-complexes is largely unexplored. Here we show that Arp3B, but not Arp3 is subject to regulation by the methionine monooxygenase MICAL2, which is recruited to branched actin networks by coronin-1C. Although Arp3 and Arp3B iso-complexes promote actin assembly equally efficiently in vitro, they have different cellular properties. Arp3B turns over significantly faster than Arp3 within the network and upon its depletion actin turnover decreases. Substitution of Arp3B Met293 by Thr, the corresponding residue in Arp3 increases actin network stability, and conversely, replacing Arp3 Thr293 with Gln to mimic Met oxidation promotes network disassembly. Thus, MICAL2 regulates a subset of Arp2/3 complexes to control branched actin network disassembly.

scholarly journals Synergy between Wsp1 and Dip1 may initiate assembly of endocytic actin networks

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Connor J Balzer ◽  
Michael L James ◽  
Heidy Y Narvaez-Ortiz ◽  
Luke A Helgeson ◽  
Vladimir Sirotkin ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Schizosaccharomyces Pombe ◽  
Actin Filament ◽  
Single Molecule ◽  
Actin Filaments ◽  
Total Internal Reflection ◽  
Actin Assembly ◽  
Actin Network ◽  
Actin Networks ◽  
Co Activation

The actin filament nucleator Arp2/3 complex is activated at cortical sites in Schizosaccharomyces pombe to assemble branched actin networks that drive endocytosis. Arp2/3 complex activators Wsp1 and Dip1 are required for proper actin assembly at endocytic sites, but how they coordinately control Arp2/3-mediated actin assembly is unknown. Alone, Dip1 activates Arp2/3 complex without preexisting actin filaments to nucleate ‘seed’ filaments that activate Wsp1-bound Arp2/3 complex, thereby initiating branched actin network assembly. In contrast, because Wsp1 requires preexisting filaments to activate, it has been assumed to function exclusively in propagating actin networks by stimulating branching from preexisting filaments. Here we show that Wsp1 is important not only for propagation but also for initiation of endocytic actin networks. Using single molecule total internal reflection fluorescence microscopy we show that Wsp1 synergizes with Dip1 to co-activate Arp2/3 complex. Synergistic co-activation does not require preexisting actin filaments, explaining how Wsp1 contributes to actin network initiation in cells.

scholarly journals Synergy between Wsp1 and Dip1 may initiate assembly of endocytic actin networks

2020 ◽  
Author(s):  
Connor J. Balzer ◽  
Michael L. James ◽  
Luke A. Helgeson ◽  
Vladimir Sirotkin ◽  
Brad J. Nolen
Keyword(s):  
Actin Filament ◽  
Single Molecule ◽  
Actin Filaments ◽  
Actin Assembly ◽  
Actin Network ◽  
Tirf Microscopy ◽  
Actin Networks ◽  
In Cells

AbstractThe actin filament nucleator Arp2/3 complex is activated at cortical sites in S. pombe to assemble branched actin networks that drive endocytosis. Arp2/3 complex activators Wsp1 and Dip1 are required for proper actin assembly at endocytic sites, but how they coordinately control Arp2/3-mediated actin assembly is unknown. Alone, Dip1 activates Arp2/3 complex without preexisting actin filaments to nucleate “seed” filaments that activate Wsp1-bound Arp2/3 complex, thereby initiating branched actin network assembly. In contrast, because Wsp1 requires pre-existing filaments to activate, it has been assumed to function exclusively in propagating actin networks by stimulating branching from pre-existing filaments. Here we show that Wsp1 is important not only for propagation, but also for initiation of endocytic actin networks. Using single molecule TIRF microscopy we show that Wsp1 synergizes with Dip1 to co-activate Arp2/3 complex. Synergistic coactivation does not require pre-existing actin filaments, explaining how Wsp1 contributes to actin network initiation in cells.

scholarly journals Revolving movement of a dynamic cluster of actin filaments during mitosis

2010 ◽  
Vol 191 (3) ◽  
pp. 453-462 ◽  
Author(s):  
Masaru Mitsushima ◽  
Kazuhiro Aoki ◽  
Miki Ebisuya ◽  
Shigeru Matsumura ◽  
Takuya Yamamoto ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Filaments ◽  
Angular Speed ◽  
Cell Cortex ◽  
Actin Assembly ◽  
Contractile Ring ◽  
Dynamic Cluster ◽  
Rapid Changes ◽  
M Phase ◽  
Key Factor

The actin cytoskeleton undergoes rapid changes in its architecture during mitosis. Here, we demonstrate novel actin assembly dynamics in M phase. An amorphous cluster of actin filaments appears during prometaphase, revolves horizontally along the cell cortex at a constant angular speed, and fuses into the contractile ring after three to four revolutions. Cdk1 activity is required for the formation of this mitotic actin cluster and its revolving movement. Rapid turnover of actin in the filaments takes place everywhere in the cluster and is also required for its cluster rotation during mitosis. Knockdown of Arp3, a component of the actin filament–nucleating Arp2/3 complex, inhibits the formation of the mitotic actin cluster without affecting other actin structures. These results identify Arp2/3 complex as a key factor in the generation of the dynamic actin cluster during mitosis.

scholarly journals Coordinated regulation of platelet actin filament barbed ends by gelsolin and capping protein.

1996 ◽  
Vol 134 (2) ◽  
pp. 389-399 ◽  
Author(s):  
K Barkalow ◽  
W Witke ◽  
D J Kwiatkowski ◽  
J H Hartwig
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Filament ◽  
Actin Polymerization ◽  
Residual Activity ◽  
Human Platelets ◽  
Actin Assembly ◽  
Capping Protein ◽  
Capping Proteins ◽  
End Capping ◽  
Human And Mouse

Exposure of cryptic actin filament fast growing ends (barbed ends) initiates actin polymerization in stimulated human and mouse platelets. Gelsolin amplifies platelet actin assembly by severing F-actin and increasing the number of barbed ends. Actin filaments in stimulated platelets from transgenic gelsolin-null mice elongate their actin without severing. F-actin barbed end capping activity persists in human platelet extracts, depleted of gelsolin, and the heterodimeric capping protein (CP) accounts for this residual activity. 35% of the approximately 5 microM CP is associated with the insoluble actin cytoskeleton of the resting platelet. Since resting platelets have an F-actin barbed end concentration of approximately 0.5 microM, sufficient CP is bound to cap these ends. CP is released from OG-permeabilized platelets by treatment with phosphatidylinositol 4,5-bisphosphate or through activation of the thrombin receptor. However, the fraction of CP bound to the actin cytoskeleton of thrombin-stimulated mouse and human platelets increases rapidly to approximately 60% within 30 s. In resting platelets from transgenic mice lacking gelsolin, which have 33% more F-actin than gelsolin-positive cells, there is a corresponding increase in the amount of CP associated with the resting cytoskeleton but no change with stimulation. These findings demonstrate an interaction between the two major F-actin barbed end capping proteins of the platelet: gelsolin-dependent severing produces barbed ends that are capped by CP. Phosphatidylinositol 4,5-bisphosphate release of gelsolin and CP from platelet cytoskeleton provides a mechanism for mediating barbed end exposure. After actin assembly, CP reassociates with the new actin cytoskeleton.

scholarly journals Cooperation between tropomyosin and α-actinin inhibits fimbrin association with actin filament networks in fission yeast

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Jenna R Christensen ◽  
Kaitlin E Homa ◽  
Alisha N Morganthaler ◽  
Rachel R Brown ◽  
Cristian Suarez ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Cellular Networks ◽  
Residence Time ◽  
Actin Filament ◽  
Binding Proteins ◽  
Actin Binding ◽  
Contractile Ring ◽  
Actin Networks ◽  
Filament Networks ◽  
Actin Patch

We previously discovered that competition between fission yeast actin binding proteins (ABPs) for binding F-actin facilitates their sorting to different cellular networks. Specifically, competition between endocytic actin patch ABPs fimbrin Fim1 and cofilin Adf1 enhances their activities, and prevents tropomyosin Cdc8’s association with actin patches. However, these interactions do not explain how Fim1 is prevented from associating strongly with other F-actin networks such as the contractile ring. Here, we identified α-actinin Ain1, a contractile ring ABP, as another Fim1 competitor. Fim1 competes with Ain1 for association with F-actin, which is dependent upon their F-actin residence time. While Fim1 outcompetes both Ain1 and Cdc8 individually, Cdc8 enhances the F-actin bundling activity of Ain1, allowing Ain1 to generate F-actin bundles that Cdc8 can bind in the presence of Fim1. Therefore, the combination of contractile ring ABPs Ain1 and Cdc8 is capable of inhibiting Fim1’s association with F-actin networks.

scholarly journals Rho and F-actin self-organize within an artificial cell cortex

2021 ◽  
Author(s):  
Jennifer Landino ◽  
Marcin Leda ◽  
Ani Michaud ◽  
Zachary T. Swider ◽  
Mariah Prom ◽  
...  
Keyword(s):  
Cell Division ◽  
Rho Gtpase ◽  
Cortical Excitability ◽  
Cell Cortex ◽  
List Type ◽  
Actin Assembly ◽  
Cortical Dynamics ◽  
Artificial Cell ◽  
Egg Extract ◽  
Dynamic Patterns

SummaryThe cell cortex, comprised of the plasma membrane and underlying cytoskeleton, undergoes dynamic reorganizations during a variety of essential biological processes including cell adhesion, cell migration, and cell division1,2. During cell division and cell locomotion, for example, waves of filamentous-actin (F-actin) assembly and disassembly develop in the cell cortex in a process termed “cortical excitability”3–7. In developing frog and starfish embryos, cortical excitability is generated through coupled positive and negative feedback, with rapid activation of Rho-mediated F-actin assembly followed in space and time by F-actin-dependent inhibition of Rho8,9. These feedback loops are proposed to serve as a mechanism for amplification of active Rho signaling at the cell equator to support furrowing during cytokinesis, while also maintaining flexibility for rapid error correction in response to movement of the mitotic spindle during chromosome segregation10. In this paper, we develop an artificial cortex based on Xenopus egg extract and supported lipid bilayers (SLBs), to investigate cortical Rho and F-actin dynamics11. This reconstituted system spontaneously develops two distinct dynamic patterns: singular excitable Rho and F-actin waves and non-traveling oscillatory Rho and F-actin patches. Both types of dynamic patterns have properties and dependencies similar to the cortical excitability previously characterized in vivo9. These findings directly support the longstanding speculation that the cell cortex is a self-organizing structure and present a novel approach for investigating mechanisms of Rho-GTPase-mediated cortical dynamics.HighlightsAn artificial cell cortex comprising Xenopus egg extract on a supported lipid bilayer self-organizes into complex, dynamic patterns of active Rho and F-actinWe identified two types of reconstituted cortical dynamics – excitable waves and coherent oscillationsReconstituted waves and oscillations require Rho activity and F-actin polymerization

scholarly journals Tropomyosin and α-actinin cooperation inhibits fimbrin association with actin filament networks in fission yeast

2017 ◽  
Author(s):  
Jenna R. Christensen ◽  
Kaitlin E. Homa ◽  
Meghan E. O’Connell ◽  
David R. Kovar
Keyword(s):  
Fission Yeast ◽  
Fluorescent Imaging ◽  
Actin Binding ◽  
Yeast Genetics ◽  
Actin Network ◽  
Contractile Ring ◽  
Actin Networks ◽  
Filament Networks ◽  
Actin Patch

ABSTRACTWe previously discovered that competition between fission yeast actin binding proteins (ABPs) for association with F-actin helps facilitate their sorting to different F-actin networks. Specifically, competition between actin patch ABPs fimbrin Fim1 and cofilin Adf1 enhances each other’s activities, and rapidly displaces tropomyosin Cdc8 from the F-actin network. However, these interactions don’t explain how Fim1, a robust competitor, is prevented from associating equally well with other F-actin networks. Here, with a combination of fission yeast genetics, live cell fluorescent imaging, and in vitro TIRF microscopy, we identified the contractile ring ABP α-actinin Ain1 as a key sorting factor. Fim1 competes with Ain1 for association with F-actin, which is dependent upon their residence time on F-actin. Remarkably, although Fim1 outcompetes both contractile ring ABPs Ain1 and Cdc8 individually, Cdc8 enhances the bundling activity of Ain1 10-fold, allowing the combination of Ain1 and Cdc8 to inhibit Fim1 association with contractile ring F-actin.

scholarly journals Synergy between Cyclase-associated protein and Cofilin accelerates actin filament depolymerization by two orders of magnitude

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Shashank Shekhar ◽  
Johnson Chung ◽  
Jane Kondev ◽  
Jeff Gelles ◽  
Bruce L. Goode
Keyword(s):  
Actin Filament ◽  
Single Molecule ◽  
Actin Filaments ◽  
Binding Protein ◽  
Actin Dynamics ◽  
Actin Network ◽  
Cellular Mechanisms ◽  
Actin Networks ◽  
Binding Event

AbstractCellular actin networks can be rapidly disassembled and remodeled in a few seconds, yet in vitro actin filaments depolymerize slowly over minutes. The cellular mechanisms enabling actin to depolymerize this fast have so far remained obscure. Using microfluidics-assisted TIRF, we show that Cyclase-associated protein (CAP) and Cofilin synergize to processively depolymerize actin filament pointed ends at a rate 330-fold faster than spontaneous depolymerization. Single molecule imaging further reveals that hexameric CAP molecules interact with the pointed ends of Cofilin-decorated filaments for several seconds at a time, removing approximately 100 actin subunits per binding event. These findings establish a paradigm, in which a filament end-binding protein and a side-binding protein work in concert to control actin dynamics, and help explain how rapid actin network depolymerization is achieved in cells.

Regulation of Actin Assembly Associated With Protrusion and Adhesion in Cell Migration

2008 ◽  
Vol 88 (2) ◽  
pp. 489-513 ◽  
Author(s):  
Christophe Le Clainche ◽  
Marie-France Carlier
Keyword(s):  
Cell Migration ◽  
Actin Cytoskeleton ◽  
Actin Filaments ◽  
Actin Assembly ◽  
Concerted Action ◽  
Actin Networks ◽  
Capping Proteins ◽  
Cell Matrix ◽  
A Cell ◽  
Actomyosin Contraction

To migrate, a cell first extends protrusions such as lamellipodia and filopodia, forms adhesions, and finally retracts its tail. The actin cytoskeleton plays a major role in this process. The first part of this review (sect. ii) describes the formation of the lamellipodial and filopodial actin networks. In lamellipodia, the WASP-Arp2/3 pathways generate a branched filament array. This polarized dendritic actin array is maintained in rapid treadmilling by the concerted action of ADF, profilin, and capping proteins. In filopodia, formins catalyze the processive assembly of nonbranched actin filaments. Cell matrix adhesions mechanically couple actin filaments to the substrate to convert the treadmilling into protrusion and the actomyosin contraction into traction of the cell body and retraction of the tail. The second part of this review (sect. iii) focuses on the function and the regulation of major proteins (vinculin, talin, tensin, and α-actinin) that control the nucleation, the binding, and the barbed-end growth of actin filaments in adhesions.

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