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 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 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 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 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.

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 Characterization of structural and functional domains of the anillin-related protein Mid1p that contribute to cytokinesis in fission yeast

2012 ◽  
Vol 23 (20) ◽  
pp. 3993-4007 ◽  
Author(s):  
Shambaditya Saha ◽  
Thomas D. Pollard
Keyword(s):  
Fission Yeast ◽  
Myosin Ii ◽  
Full Length ◽  
Yeast Cells ◽  
Related Protein ◽  
Contractile Ring ◽  
Structural Domains ◽  
Biophysical Analysis ◽  
Direct Condensation

Fission yeast cells depend on the anillin-related protein Mid1p for reliable cytokinesis. Insolubility limits the purification of full-length Mid1p for biophysical analysis, and lack of knowledge about the structural domains of Mid1p limits functional analysis. We addressed these limitations by identifying in a bacterial expression screen of random Mid1p fragments five soluble segments that can be purified and one insoluble segment. Using complementation experiments in Δmid1 cells, we tested the biological functions of these six putative domains that account for full-length Mid1p. The N-terminal domain (residues 1–149) is essential for correct positioning and orientation of septa. The third domain (residues 309–452) allows the construct composed of the first three domains (residues 1-452) to form hydrodynamically well-behaved octamers. Constructs consisting of residues 1–452 or 1–578 carry out most functions of full-length Mid1p, including concentration at the equatorial cortex in nodes that accumulate myosin-II and other contractile ring proteins during mitosis. However, cells depending on these constructs without the insoluble domain (residues 579–797) form equatorially located rings slowly from strands rather than by direct condensation of nodes. We conclude that residues 1–578 assemble node components myosin-II, Rng2p, and Cdc15p, and the insoluble domain facilitates the normal, efficient condensation of nodes into rings.

scholarly journals Molecular organization of cytokinesis nodes and contractile rings by super-resolution fluorescence microscopy of live fission yeast

2016 ◽  
Vol 113 (40) ◽  
pp. E5876-E5885 ◽  
Author(s):  
Caroline Laplante ◽  
Fang Huang ◽  
Irene R. Tebbs ◽  
Joerg Bewersdorf ◽  
Thomas D. Pollard
Keyword(s):  
Fission Yeast ◽  
High Speed ◽  
Myosin Ii ◽  
Super Resolution ◽  
Molecular Organization ◽  
Yeast Cells ◽  
Contractile Ring ◽  
Actin Ring ◽  
Structural Units ◽  
Iq Motif

Cytokinesis in animals, fungi, and amoebas depends on the constriction of a contractile ring built from a common set of conserved proteins. Many fundamental questions remain about how these proteins organize to generate the necessary tension for cytokinesis. Using quantitative high-speed fluorescence photoactivation localization microscopy (FPALM), we probed this question in live fission yeast cells at unprecedented resolution. We show that nodes, protein assembly precursors to the contractile ring, are discrete structural units with stoichiometric ratios and distinct distributions of constituent proteins. Anillin Mid1p, Fes/CIP4 homology-Bin/amphiphysin/Rvs (F-BAR) Cdc15p, IQ motif containing GTPase-activating protein (IQGAP) Rng2p, and formin Cdc12p form the base of the node that anchors the ends of myosin II tails to the plasma membrane, with myosin II heads extending into the cytoplasm. This general node organization persists in the contractile ring where nodes move bidirectionally during constriction. We observed the dynamics of the actin network during cytokinesis, starting with the extension of short actin strands from nodes, which sometimes connected neighboring nodes. Later in cytokinesis, a broad network of thick bundles coalesced into a tight ring around the equator of the cell. The actin ring was ∼125 nm wide and ∼125 nm thick. These observations establish the organization of the proteins in the functional units of a cytokinetic contractile ring.

scholarly journals Myosin-II reorganization during mitosis is controlled temporally by its dephosphorylation and spatially by Mid1 in fission yeast

2004 ◽  
Vol 165 (5) ◽  
pp. 685-695 ◽  
Author(s):  
Fumio Motegi ◽  
Mithilesh Mishra ◽  
Mohan K. Balasubramanian ◽  
Issei Mabuchi
Keyword(s):  
Fission Yeast ◽  
Myosin Ii ◽  
Phosphorylation Site ◽  
Terminal Region ◽  
Contractile Ring ◽  
Actin Ring ◽  
Division Site ◽  
Ring Assembly ◽  
Medial Cortex ◽  
Early Mitosis

Cytokinesis in many eukaryotes requires an actomyosin contractile ring. Here, we show that in fission yeast the myosin-II heavy chain Myo2 initially accumulates at the division site via its COOH-terminal 134 amino acids independently of F-actin. The COOH-terminal region can access to the division site at early G2, whereas intact Myo2 does so at early mitosis. Ser1444 in the Myo2 COOH-terminal region is a phosphorylation site that is dephosphorylated during early mitosis. Myo2 S1444A prematurely accumulates at the future division site and promotes formation of an F-actin ring even during interphase. The accumulation of Myo2 requires the anillin homologue Mid1 that functions in proper ring placement. Myo2 interacts with Mid1 in cell lysates, and this interaction is inhibited by an S1444D mutation in Myo2. Our results suggest that dephosphorylation of Myo2 liberates the COOH-terminal region from an intramolecular inhibition. Subsequently, dephosphorylated Myo2 is anchored by Mid1 at the medial cortex and promotes the ring assembly in cooperation with F-actin.

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