scholarly journals Assembly of the cytokinetic contractile ring from a broad band of nodes in fission yeast

2006 ◽  
Vol 174 (3) ◽  
pp. 391-402 ◽  
Author(s):  
Jian-Qiu Wu ◽  
Vladimir Sirotkin ◽  
David R. Kovar ◽  
Matthew Lord ◽  
Christopher C. Beltzner ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Fusion Proteins ◽  
Cell Separation ◽  
Actin Polymerization ◽  
Myosin Ii ◽  
Broad Band ◽  
Compact Ring ◽  
Contractile Ring ◽  
Single Spot ◽  
Anaphase B

We observed live fission yeast expressing pairs of functional fluorescent fusion proteins to test the popular model that the cytokinetic contractile ring assembles from a single myosin II progenitor or a Cdc12p-Cdc15p spot. Under our conditions, the anillin-like protein Mid1p establishes a broad band of small dots or nodes in the cortex near the nucleus. These nodes mature by the addition of conventional myosin II (Myo2p, Cdc4p, and Rlc1p), IQGAP (Rng2p), pombe Cdc15 homology protein (Cdc15p), and formin (Cdc12p). The nodes coalesce laterally into a compact ring when Cdc12p and profilin Cdc3p stimulate actin polymerization. We did not observe assembly of contractile rings by extension of a leading cable from a single spot or progenitor. Arp2/3 complex and its activators accumulate in patches near the contractile ring early in anaphase B, but are not concentrated in the contractile ring and are not required for assembly of the contractile ring. Their absence delays late steps in cytokinesis, including septum formation and cell separation.

scholarly journals Myosins generate contractile force and maintain organization in the cytokinetic contractile ring

2021 ◽  
Author(s):  
Zachary A. McDargh ◽  
Shuyuan Wang ◽  
Harvey F. Chin ◽  
Sathish Thiyagarajan ◽  
Erdem Karatekin ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Striated Muscle ◽  
Protein Complexes ◽  
Myosin Ii ◽  
Force Production ◽  
Super Resolution ◽  
Building Blocks ◽  
Contractile Ring ◽  
Ring Model ◽  
Self Assembling

During cytokinesis, cells assemble an actomyosin contractile ring whose tension constricts and divides cells, but the ring tension was rarely measured. Actomyosin force generation is well understood for the regular sarcomeric architecture of striated muscle, but recent super-resolution studies of fission yeast contractile rings revealed organizational building blocks that are not sarcomeres but irregularly positioned plasma membrane-anchored protein complexes called nodes. Here, we measured contractile ring tensions in fission yeast protoplast cells. The myosin II isoforms Myo2 and Myp2 generated the tension, with a ~2-fold greater contribution from Myo2. Simulations of a molecularly detailed ring model revealed a sliding node mechanism for tension, where nodes hosting tense actin filaments were pulled bidirectionally around the ring. Myo2 and Myp2 chaperoned self-assembling components into the ring organization, and anchored the ring against bridging instabilities. Thus, beyond force production, Myo2 and Myp2 are the principal organizers, bundlers and anchors of the contractile ring.

scholarly journals α-Actinin and fimbrin cooperate with myosin II to organize actomyosin bundles during contractile-ring assembly

2012 ◽  
Vol 23 (16) ◽  
pp. 3094-3110 ◽  
Author(s):  
Damien Laporte ◽  
Nikola Ojkic ◽  
Dimitrios Vavylonis ◽  
Jian-Qiu Wu
Keyword(s):  
Actin Filaments ◽  
Myosin Ii ◽  
Broad Band ◽  
Live Cells ◽  
Condensation Process ◽  
Contractile Ring ◽  
Ring Assembly ◽  
Cooperative Process ◽  
Normal Ring ◽  
Work Supports

The actomyosin contractile ring assembles through the condensation of a broad band of nodes that forms at the cell equator in fission yeast cytokinesis. The condensation process depends on actin filaments that interconnect nodes. By mutating or titrating actin cross-linkers α-actinin Ain1 and fimbrin Fim1 in live cells, we reveal that both proteins are involved in node condensation. Ain1 and Fim1 stabilize the actin cytoskeleton and modulate node movement, which prevents nodes and linear structures from aggregating into clumps and allows normal ring formation. Our computer simulations modeling actin filaments as semiflexible polymers reproduce the experimental observations and provide a model of how actin cross-linkers work with other proteins to regulate actin-filament orientations inside actin bundles and organize the actin network. As predicted by the simulations, doubling myosin II Myo2 level rescues the node condensation defects caused by Ain1 overexpression. Taken together, our work supports a cooperative process of ring self-organization driven by the interaction between actin filaments and myosin II, which is progressively stabilized by the cross-linking proteins.

scholarly journals Tropomyosin and Myosin-II Cellular Levels Promote Actomyosin Ring Assembly in Fission Yeast

2010 ◽  
Vol 21 (6) ◽  
pp. 989-1000 ◽  
Author(s):  
Benjamin C. Stark ◽  
Thomas E. Sladewski ◽  
Luther W. Pollard ◽  
Matthew Lord
Keyword(s):  
Motor Activity ◽  
Atpase Activity ◽  
Fission Yeast ◽  
Actin Filaments ◽  
Myosin Ii ◽  
Bound State ◽  
Contractile Ring ◽  
Ring Assembly

Myosin-II (Myo2p) and tropomyosin are essential for contractile ring formation and cytokinesis in fission yeast. Here we used a combination of in vivo and in vitro approaches to understand how these proteins function at contractile rings. We find that ring assembly is delayed in Myo2p motor and tropomyosin mutants, but occurs prematurely in cells engineered to express two copies of myo2. Thus, the timing of ring assembly responds to changes in Myo2p cellular levels and motor activity, and the emergence of tropomyosin-bound actin filaments. Doubling Myo2p levels suppresses defects in ring assembly associated with a tropomyosin mutant, suggesting a role for tropomyosin in maximizing Myo2p function. Correspondingly, tropomyosin increases Myo2p actin affinity and ATPase activity and promotes Myo2p-driven actin filament gliding in motility assays. Tropomyosin achieves this by favoring the strong actin-bound state of Myo2p. This mode of regulation reflects a role for tropomyosin in specifying and stabilizing actomyosin interactions, which facilitates contractile ring assembly in the fission yeast system.

scholarly journals Fission yeast IQGAP maintains F-actin-independent localization of myosin-II in the contractile ring

2013 ◽  
Vol 19 (2) ◽  
pp. 161-176 ◽  
Author(s):  
Masak Takaine ◽  
Osamu Numata ◽  
Kentaro Nakano
Keyword(s):  
Fission Yeast ◽  
Myosin Ii ◽  
Contractile Ring

scholarly journals Mid2p stabilizes septin rings during cytokinesis in fission yeast

2003 ◽  
Vol 160 (7) ◽  
pp. 1083-1092 ◽  
Author(s):  
Ana Berlin ◽  
Anne Paoletti ◽  
Fred Chang
Keyword(s):  
Fission Yeast ◽  
Cell Separation ◽  
Sequence Similarity ◽  
Ring Closure ◽  
Pleckstrin Homology Domain ◽  
Wild Type ◽  
Contractile Ring ◽  
Single Ring ◽  
And Function ◽  
Cell Cell

Septins are filament-forming proteins with a conserved role in cytokinesis. In the fission yeast Schizosaccharomyces pombe, septin rings appear to be involved primarily in cell–cell separation, a late stage in cytokinesis. Here, we identified a protein Mid2p on the basis of its sequence similarity to S. pombe Mid1p, Saccharomyces cerevisiae Bud4p, and Candida albicans Int1p. Like septin mutants, mid2Δ mutants had delays in cell–cell separation. mid2Δ mutants were defective in septin organization but not contractile ring closure or septum formation. In wild-type cells, septins assembled first during mitosis in a single ring and during septation developed into double rings that did not contract. In mid2Δ cells, septins initially assembled in a single ring but during septation appeared in the cleavage furrow, forming a washer or disc structure. FRAP studies showed that septins are stable in wild-type cells but exchange 30-fold more rapidly in mid2Δ cells. Mid2p colocalized with septins and required septins for its localization. A COOH-terminal pleckstrin homology domain of Mid2p was required for its localization and function. No genetic interactions were found between mid2 and the related gene mid1. Thus, these studies identify a new factor responsible for the proper stability and function of septins during cytokinesis.

scholarly journals Cytokinesis Depends on the Motor Domains of Myosin-II in Fission Yeast but Not in Budding Yeast

2005 ◽  
Vol 16 (11) ◽  
pp. 5346-5355 ◽  
Author(s):  
Matthew Lord ◽  
Ellen Laves ◽  
Thomas D. Pollard
Keyword(s):  
Atpase Activity ◽  
Fission Yeast ◽  
Mechanism Of Action ◽  
Budding Yeast ◽  
Myosin Ii ◽  
Cellular Function ◽  
Motor Domain ◽  
Actin Binding ◽  
Contractile Ring ◽  
Head Domain

Budding yeast possesses one myosin-II, Myo1p, whereas fission yeast has two, Myo2p and Myp2p, all of which contribute to cytokinesis. We find that chimeras consisting of Myo2p or Myp2p motor domains fused to the tail of Myo1p are fully functional in supporting budding yeast cytokinesis. Remarkably, the tail alone of budding yeast Myo1p localizes to the contractile ring, supporting both its constriction and cytokinesis. In contrast, fission yeast Myo2p and Myp2p require both the catalytic head domain as well as tail domains for function, with the tails providing distinct functions ( Bezanilla and Pollard, 2000 ). Myo1p is the first example of a myosin whose cellular function does not require a catalytic motor domain revealing a novel mechanism of action for budding yeast myosin-II independent of actin binding and ATPase activity.

scholarly journals Fission yeast TRP channel Pkd2p localizes to the cleavage furrow and regulates cell separation during cytokinesis

2019 ◽  
Vol 30 (15) ◽  
pp. 1791-1804 ◽  
Author(s):  
Zachary Morris ◽  
Debatrayee Sinha ◽  
Abhishek Poddar ◽  
Brittni Morris ◽  
Qian Chen
Keyword(s):  
Fission Yeast ◽  
Cell Separation ◽  
Secretory Pathway ◽  
Transient Receptor Potential ◽  
Turgor Pressure ◽  
Receptor Potential ◽  
Mitogen Activated Protein Kinase ◽  
Force Sensing ◽  
Cleavage Furrow ◽  
Contractile Ring

Force plays a central role in separating daughter cells during cytokinesis, the last stage of cell division. However, the mechanism of force sensing during cytokinesis remains unknown. Here we discovered that Pkd2p, a putative force-sensing transient receptor potential channel, localizes to the cleavage furrow during cytokinesis of the fission yeast, Schizosaccharomyces pombe. Pkd2p, whose human homologues are associated with autosomal polycystic kidney disease, is an essential protein whose localization depends on the contractile ring and the secretory pathway. We identified and characterized a novel pkd2 mutant pkd2-81KD. The pkd2 mutant cells show signs of osmotic stress, including temporary shrinking, paused turnover of the cytoskeletal structures, and hyperactivated mitogen-activated protein kinase signaling. During cytokinesis, although the contractile ring constricts more rapidly in the pkd2 mutant than the wild-type cells (50% higher), the cell separation in the mutant is slower and often incomplete. These cytokinesis defects are also consistent with misregulated turgor pressure. Finally, the pkd2 mutant exhibits strong genetic interactions with two mutants of the septation initiation network pathway, a signaling cascade essential for cytokinesis. We propose that Pkd2p modulates osmotic homeostasis and is potentially a novel regulator of cytokinesis.

scholarly journals Role of Tropomyosin in Formin-mediated Contractile Ring Assembly in Fission Yeast

2009 ◽  
Vol 20 (8) ◽  
pp. 2160-2173 ◽  
Author(s):  
Colleen T. Skau ◽  
Erin M. Neidt ◽  
David R. Kovar
Keyword(s):  
Fission Yeast ◽  
Actin Filament ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Actin Assembly ◽  
Animal Cells ◽  
Contractile Ring ◽  
Direct Role ◽  
Ring Assembly

Like animal cells, fission yeast divides by assembling actin filaments into a contractile ring. In addition to formin Cdc12p and profilin, the single tropomyosin isoform SpTm is required for contractile ring assembly. Cdc12p nucleates actin filaments and remains processively associated with the elongating barbed end while driving the addition of profilin-actin. SpTm is thought to stabilize mature filaments, but it is not known how SpTm localizes to the contractile ring and whether SpTm plays a direct role in Cdc12p-mediated actin polymerization. Using “bulk” and single actin filament assays, we discovered that Cdc12p can recruit SpTm to actin filaments and that SpTm has diverse effects on Cdc12p-mediated actin assembly. On its own, SpTm inhibits actin filament elongation and depolymerization. However, Cdc12p completely overcomes the combined inhibition of actin nucleation and barbed end elongation by profilin and SpTm. Furthermore, SpTm increases the length of Cdc12p-nucleated actin filaments by enhancing the elongation rate twofold and by allowing them to anneal end to end. In contrast, SpTm ultimately turns off Cdc12p-mediated elongation by “trapping” Cdc12p within annealed filaments or by dissociating Cdc12p from the barbed end. Therefore, SpTm makes multiple contributions to contractile ring assembly during and after actin polymerization.

scholarly journals Progress towards understanding the mechanism of cytokinesis in fission yeast

2008 ◽  
Vol 36 (3) ◽  
pp. 425-430 ◽  
Author(s):  
Thomas D. Pollard
Keyword(s):  
Fission Yeast ◽  
Actin Filaments ◽  
Biochemical Analysis ◽  
Fluorescent Proteins ◽  
Myosin Ii ◽  
Live Cells ◽  
Contractile Ring ◽  
Simple Hypothesis ◽  
A Cell ◽  
Temporal And Spatial

We use fission yeast to study the molecular mechanism of cytokinesis. We benefit from a long history in genetic analysis of the cell cycle in fission yeast, which provided the most complete inventory of cytokinesis proteins. We used fluorescence microscopy of proteins tagged with fluorescent proteins to establish the temporal and spatial pathway for the assembly and constriction of the contractile ring. We combined biochemical analysis of purified proteins (myosin-II, profilin, formin Cdc12p and cofilin), observations of fluorescent fusion proteins in live cells and mathematical modelling to formulate and test a simple hypothesis for the assembly of the contractile ring. This model involves the formation of 65 nodes containing myosin-II and formin Cdc12p around the equator of the cell. As a cell enters anaphase, actin filaments grow from formin Cdc12p in these nodes. Myosin captures actin filaments from adjacent nodes and pulls intermittently to condense the nodes into a contractile ring.

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