The formins Cdc12 and For3 cooperate during contractile ring assembly in cytokinesis

Coffman, Valerie C.; Sees, Jennifer A.; Kovar, David R.; Wu, Jian-Qiu

doi:10.1083/jcb.201305022

Cited by 47 publications

(78 citation statements)

References 101 publications

(182 reference statements)

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“…Much of the actin in the ring is either Cdc12p-derived or For3-derived (36,37). It has been proposed that actin filaments might remain engaged with nodebound Cdc12p at their barbed ends, thus connecting filaments to the membrane (14,30).…”

Section: Discussionmentioning

confidence: 99%

Structure of the fission yeast actomyosin ring during constriction

Swulius

Nguyen

Ladinsky

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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Cell division in many eukaryotes is driven by a ring containing actin and myosin. While much is known about the main proteins involved, the precise arrangement of actin filaments within the contractile machinery, and how force is transmitted to the membrane, remains unclear. Here we use cryosectioning and cryofocused ion beam milling to gain access to cryopreserved actomyosin rings in Schizosaccharomyces pombe for direct 3D imaging by electron cryotomography. Our results show that straight, overlapping actin filaments, running nearly parallel to each other and to the membrane, form a loose bundle of ∼150 nm in diameter that "saddles" the inward-bending membrane at the leading edge of the division septum. The filaments do not make direct contact with the membrane. Our analysis of the actin filaments reveals the variability in filament number, nearest-neighbor distances between filaments within the bundle, their distance from the membrane, and angular distribution with respect to the membrane.C ytokinesis, the final step of cell division in eukaryotic cells, is typically driven by a contractile actomyosin ring (AMR) primarily composed of actin (1) and myosin (2). Our understanding of the molecular mechanisms of cytokinesis is most detailed in the rodshaped unicellular eukaryote Schizosaccharomyces pombe (otherwise known as fission yeast), which shares a remarkably conserved set of cytokinesis genes with metazoans (3). In S. pombe, the AMR undergoes multiple phases known as assembly, maturation, constriction, and disassembly (4), with open questions in each of these four stages. Due to a lack of information about the precise arrangement of filamentous actin (F-actin) within the force-generating network of the AMR, we chose to focus on imaging the AMR during constriction.In S. pombe, glancing sections through plastic-embedded, dividing cells gave the first glimpse of actin filaments running parallel to the division plane at the front of the septum (5). Unfortunately, the study yielded limited examples and lacked 3D information for a full analysis. In an ambitious pioneering effort, Kamasaki et al. (6) produced 3D reconstructions of entire S. pombe AMRs by imaging serial sections through permeabilized cells decorated with myosin S1 fragments. The amount of F-actin and the size of the rings appeared significantly altered by the procedure used for preserving them (details in Discussion), but the continuous bundles that were reconstructed were composed of mixed polarity filaments running circumferentially around the cell.Here we sought to visualize the precise arrangement of F-actin within the AMR and its interface with the membrane by imaging intact cells in a cryopreserved state using electron cryotomography (ECT) (7). Because whole S. pombe cells are too thick for ECT, which is limited to specimens thinner than a few hundred nanometers, we overcame this obstacle by first rapid freezing dividing cells and then either cryosectioning them or using the recently developed method cryofocused ion beam (cryo-FIB) milling to ...

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Section: Discussionmentioning

confidence: 99%

Structure of the fission yeast actomyosin ring during constriction

Swulius

Nguyen

Ladinsky

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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“…Gef1-mNG persists on both sides of the septum barrier in 40% of cells treated with LatA, but not in mock DMSO-treated cells ( Figure 3C). Cells undergoing ring constriction and septum formation display actin cables as well as Arp2/3-complex-dependent patches at the division site (Coffman et al, 2013;Gachet and Hyams, 2005;Huang et al, 2012;Wang et al, 2016). LatA treatment removes all types of filamentous actin structures (Spector et al, 1983).…”

Section: Scd1 Promotes Gef1 Removal From the Division Site At The Endmentioning

confidence: 99%

A novel interplay between GEFs orchestrates Cdc42 activity during cell polarity and cytokinesis

Hercyk

Rich

Das

2018

Preprint

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Cdc42, a conserved regulator of cell polarity, is activated by two GEFs, Gef1 and Scd1, in fission yeast. While gef1 and scd1 mutants exhibit distinct phenotypes, how they do so is unclear given that they activate the same GTPase. Using the GEF localization pattern during cytokinesis as a paradigm, we report a novel interplay between Gef1 and Scd1 that spatially modulates Cdc42. We find that Gef1 promotes Scd1 localization to the division site during cytokinesis and to the new end during polarized growth through the recruitment of the scaffold Scd2 via a Cdc42 feedforward pathway. Gef1-mediated Scd1 recruitment at the new end enables the transition from monopolar to bipolar growth. Reciprocally, Scd1 restricts Gef1 localization to prevent ectopic Cdc42 activation during cytokinesis to promote cell separation and during interphase to maintain cell shape. Our findings reveal an elegant regulatory pattern in which Gef1 establishes new sites of Scd1-mediated Cdc42 activity, while Scd1 restricts Gef1 to functional sites. We propose that crosstalk between GEFs is a conserved mechanism that orchestrates Cdc42 activation during complex cellular processes.Summary StatementCdc42 GEFs Gef1 and Scd1 crosstalk to fine-tune Cdc42 activity. This crosstalk promotes bipolar growth and maintains cell shape in fission yeast.

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“…For3p binds to activated Rho3p and Cdc42p, both of which are involved in membrane trafficking (Estravis et al, 2011;Martin et al, 2007;Nakano et al, 2002), and Rho3p localizes to the Golgi and endosomes, as well as to the division site (Yu et al, 2013). Furthermore, the synthetic lethal genetic interactions of both Δrga7 and Δfor3 with mutations of the genes for myosin-II essential light chain cdc4 + and IQGAP rng2 + (Table 1) (Coffman et al, 2013) are likely to result from the combined defects in the contractile ring assembly and septum assembly. Indirect effects might delay the delivery of Rga7p to the plasma membrane in Δfor3 cells, but actin filaments polymerized by For3p could help to sort membrane cargo in the Golgi complex or serve as tracks for post-Golgi vesicles.…”

Section: Bgs4p Targeting To the Cleavage Furrow Depends On Actin Filamentioning

confidence: 99%

“…For3p assembles actin filament cables during interphase (Feierbach and Chang, 2001;Martin and Chang, 2006) but also localizes to the cleavage furrow where it contributes to the timely assembly and constriction of contractile rings (Coffman et al, 2013). For3p binds to activated Rho3p and Cdc42p, both of which are involved in membrane trafficking (Estravis et al, 2011;Martin et al, 2007;Nakano et al, 2002), and Rho3p localizes to the Golgi and endosomes, as well as to the division site (Yu et al, 2013).…”

Section: Bgs4p Targeting To the Cleavage Furrow Depends On Actin Filamentioning

confidence: 99%

A role for F-BAR protein Rga7p during cytokinesis in S. pombe

Arasada

Pollard

2015

Journal of Cell Science

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F-BAR proteins are known to participate in cytokinesis, but their mechanisms are not well understood. Here we investigated Rga7p an S. pombe F-BAR protein with a RhoGAP domain. Localization of Rga7p to the cytokinetic cleavage furrow depends on its F-BAR domain, actin filaments, formins Cdc12p and For3p, and the presence of a contractile ring. Rga7p is not required for the constriction of the contractile ring but does participate in the transport of a β-glucan synthetase Bgs4p from the late Golgi compartments to the plasma membrane adjacent to the contractile ring. Cells without Rga7p moved Bgs4p normally from the poles to the Golgi apparatus near the cell center, but Bgs4p then moved slowly from the late Golgi compartments to the cleavage site. The late arrival and lower than normal numbers of Bgs4p result in septal defects late in cytokinesis and lysis of separating cells similar to cells with the mutations in the cwg1+ gene encoding Bgs4p.

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The formins Cdc12 and For3 cooperate during contractile ring assembly in cytokinesis

Abstract: Multiple formins cooperate during cytokinesis, but their functions in de novo actin assembly at the division site play the primary role in contractile ring assembly.

Cited by 47 publications

References 101 publications

Structure of the fission yeast actomyosin ring during constriction

Structure of the fission yeast actomyosin ring during constriction

A novel interplay between GEFs orchestrates Cdc42 activity during cell polarity and cytokinesis

A role for F-BAR protein Rga7p during cytokinesis in S. pombe

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