scholarly journals Role of the Hof1–Cyk3 interaction in cleavage-furrow ingression and primary-septum formation during yeast cytokinesis

2018 ◽  
Vol 29 (5) ◽  
pp. 597-609 ◽  
Author(s):  
Meng Wang ◽  
Ryuichi Nishihama ◽  
Masayuki Onishi ◽  
John R. Pringle
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Functional Significance ◽  
Normal Rate ◽  
Cleavage Furrow ◽  
Type Ii ◽  
Septum Formation ◽  
Primary Septum ◽  
Direct Binding ◽  
Type Ii Myosin

In Saccharomyces cerevisiae, it is well established that Hof1, Cyk3, and Inn1 contribute to septum formation and cytokinesis. Because hof1∆ and cyk3∆ single mutants have relatively mild defects but hof1∆ cyk3∆ double mutants are nearly dead, it has been hypothesized that these proteins contribute to parallel pathways. However, there is also evidence that they interact physically. In this study, we examined this interaction and its functional significance in detail. Our data indicate that the interaction 1) is mediated by a direct binding of the Hof1 SH3 domain to a proline-rich motif in Cyk3; 2) occurs specifically at the time of cytokinesis but is independent of the (hyper)phosphorylation of both proteins that occurs at about the same time; 3) is dispensable for the normal localization of both proteins; 4) is essential for normal primary-septum formation and a normal rate of cleavage-furrow ingression; and 5) becomes critical for growth when either Inn1 or the type II myosin Myo1 (a key component of the contractile actomyosin ring) is absent. The similarity in phenotype between cyk3∆ mutants and mutants specifically lacking the Hof1–Cyk3 interaction suggests that the interaction is particularly important for Cyk3 function, but it may be important for Hof1 function as well.

scholarly journals Role of Inn1 and its interactions with Hof1 and Cyk3 in promoting cleavage furrow and septum formation in S. cerevisiae

2009 ◽  
Vol 185 (6) ◽  
pp. 995-1012 ◽  
Author(s):  
Ryuichi Nishihama ◽  
Jennifer H. Schreiter ◽  
Masayuki Onishi ◽  
Elizabeth A. Vallen ◽  
Julia Hanna ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Exit ◽  
Chitin Synthase ◽  
Cleavage Furrow ◽  
Sh3 Domains ◽  
C2 Domains ◽  
Division Site ◽  
Primary Septum ◽  
N Terminus

Cytokinesis requires coordination of actomyosin ring (AMR) contraction with rearrangements of the plasma membrane and extracellular matrix. In Saccharomyces cerevisiae, new membrane, the chitin synthase Chs2 (which forms the primary septum [PS]), and the protein Inn1 are all delivered to the division site upon mitotic exit even when the AMR is absent. Inn1 is essential for PS formation but not for Chs2 localization. The Inn1 C-terminal region is necessary for localization, and distinct PXXP motifs in this region mediate functionally important interactions with SH3 domains in the cytokinesis proteins Hof1 (an F-BAR protein) and Cyk3 (whose overexpression can restore PS formation in inn1Δ cells). The Inn1 N terminus resembles C2 domains but does not appear to bind phospholipids; nonetheless, when overexpressed or fused to Hof1, it can provide Inn1 function even in the absence of the AMR. Thus, Inn1 and Cyk3 appear to cooperate in activating Chs2 for PS formation, which allows coordination of AMR contraction with ingression of the cleavage furrow.

scholarly journals Involvement of an Actomyosin Contractile Ring in Saccharomyces cerevisiae Cytokinesis

1998 ◽  
Vol 142 (5) ◽  
pp. 1301-1312 ◽  
Author(s):  
Erfei Bi ◽  
Paul Maddox ◽  
Daniel J. Lew ◽  
E.D. Salmon ◽  
John N. McMillan ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Separation ◽  
Cortical Actin ◽  
Primary Defect ◽  
Septum Formation ◽  
Bud Emergence ◽  
Bud Neck ◽  
Bud Site ◽  
Type Ii Myosin

In Saccharomyces cerevisiae, the mother cell and bud are connected by a narrow neck. The mechanism by which this neck is closed during cytokinesis has been unclear. Here we report on the role of a contractile actomyosin ring in this process. Myo1p (the only type II myosin in S. cerevisiae) forms a ring at the presumptive bud site shortly before bud emergence. Myo1p ring formation depends on the septins but not on F-actin, and preexisting Myo1p rings are stable when F-actin is depolymerized. The Myo1p ring remains in the mother–bud neck until the end of anaphase, when a ring of F-actin forms in association with it. The actomyosin ring then contracts to a point and disappears. In the absence of F-actin, the Myo1p ring does not contract. After ring contraction, cortical actin patches congregate at the mother–bud neck, and septum formation and cell separation rapidly ensue. Strains deleted for MYO1 are viable; they fail to form the actin ring but show apparently normal congregation of actin patches at the neck. Some myo1Δ strains divide nearly as efficiently as wild type; other myo1Δ strains divide less efficiently, but it is unclear whether the primary defect is in cytokinesis, septum formation, or cell separation. Even cells lacking F-actin can divide, although in this case division is considerably delayed. Thus, the contractile actomyosin ring is not essential for cytokinesis in S. cerevisiae. In its absence, cytokinesis can still be completed by a process (possibly localized cell–wall synthesis leading to septum formation) that appears to require septin function and to be facilitated by F-actin.

Increased Chitin Synthesis in Response to Type II Myosin Deficiency in Saccharomyces cerevisiae

2000 ◽  
Vol 3 (1) ◽  
pp. 20-25 ◽  
Author(s):  
J.A. Cruz ◽  
R. Garcia ◽  
J.F. Rodriguez-Orengo ◽  
J.R. Rodriguez-Medina
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Type Ii ◽  
Chitin Synthesis ◽  
Type Ii Myosin

scholarly journals The Role of Calmodulin vs. Synaptotagmin in Exocytosis

2021 ◽  
Vol 14 ◽  
Author(s):  
Renhao Xue ◽  
Hao Meng ◽  
Jiaxiang Yin ◽  
Jingyao Xia ◽  
Zhitao Hu ◽  
...  
Keyword(s):  
Regulatory Proteins ◽  
Type Ii ◽  
Myosin V ◽  
Dependent Protein Kinase ◽  
High Affinity ◽  
Ef Hand ◽  
Synaptotagmin 1 ◽  
Dependent Protein ◽  
Direct Binding

Exocytosis is a Ca2+-regulated process that requires the participation of Ca2+ sensors. In the 1980s, two classes of Ca2+-binding proteins were proposed as putative Ca2+ sensors: EF-hand protein calmodulin, and the C2 domain protein synaptotagmin. In the next few decades, numerous studies determined that in the final stage of membrane fusion triggered by a micromolar boost in the level of Ca2+, the low affinity Ca2+-binding protein synaptotagmin, especially synaptotagmin 1 and 2, acts as the primary Ca2+ sensor, whereas calmodulin is unlikely to be functional due to its high Ca2+ affinity. However, in the meantime emerging evidence has revealed that calmodulin is involved in the earlier exocytotic steps prior to fusion, such as vesicle trafficking, docking and priming by acting as a high affinity Ca2+ sensor activated at submicromolar level of Ca2+. Calmodulin directly interacts with multiple regulatory proteins involved in the regulation of exocytosis, including VAMP, myosin V, Munc13, synapsin, GAP43 and Rab3, and switches on key kinases, such as type II Ca2+/calmodulin-dependent protein kinase, to phosphorylate a series of exocytosis regulators, including syntaxin, synapsin, RIM and Ca2+ channels. Moreover, calmodulin interacts with synaptotagmin through either direct binding or indirect phosphorylation. In summary, calmodulin and synaptotagmin are Ca2+ sensors that play complementary roles throughout the process of exocytosis. In this review, we discuss the complementary roles that calmodulin and synaptotagmin play as Ca2+ sensors during exocytosis.

scholarly journals Cleavage-furrow formation without F-actin inChlamydomonas

2020 ◽  
Vol 117 (31) ◽  
pp. 18511-18520
Author(s):  
Masayuki Onishi ◽  
James G. Umen ◽  
Frederick R. Cross ◽  
John R. Pringle
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Green Alga ◽  
Eukaryotic Cells ◽  
Cleavage Furrow ◽  
Type Ii ◽  
Type Ii Myosin ◽  
In Cells

It is widely believed that cleavage-furrow formation during cytokinesis is driven by the contraction of a ring containing F-actin and type-II myosin. However, even in cells that have such rings, they are not always essential for furrow formation. Moreover, many taxonomically diverse eukaryotic cells divide by furrowing but have no type-II myosin, making it unlikely that an actomyosin ring drives furrowing. To explore this issue further, we have used one such organism, the green algaChlamydomonas reinhardtii. We found that although F-actin is associated with the furrow region, none of the three myosins (of types VIII and XI) is localized there. Moreover, when F-actin was eliminated through a combination of a mutation and a drug, furrows still formed and the cells divided, although somewhat less efficiently than normal. Unexpectedly, division of the largeChlamydomonaschloroplast was delayed in the cells lacking F-actin; as this organelle lies directly in the path of the cleavage furrow, this delay may explain, at least in part, the delay in cytokinesis itself. Earlier studies had shown an association of microtubules with the cleavage furrow, and we used a fluorescently tagged EB1 protein to show that microtubules are still associated with the furrows in the absence of F-actin, consistent with the possibility that the microtubules are important for furrow formation. We suggest that the actomyosin ring evolved as one way to improve the efficiency of a core process for furrow formation that was already present in ancestral eukaryotes.

scholarly journals Biphasic targeting and cleavage furrow ingression directed by the tail of a myosin II

2010 ◽  
Vol 191 (7) ◽  
pp. 1333-1350 ◽  
Author(s):  
Xiaodong Fang ◽  
Jianying Luo ◽  
Ryuichi Nishihama ◽  
Carsten Wloka ◽  
Christopher Dravis ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Trafficking ◽  
Myosin Ii ◽  
Force Generation ◽  
Extracellular Matrix Remodeling ◽  
Cleavage Furrow ◽  
Matrix Remodeling ◽  
Division Site ◽  
Targeting Signals

Cytokinesis in animal and fungal cells utilizes a contractile actomyosin ring (AMR). However, how myosin II is targeted to the division site and promotes AMR assembly, and how the AMR coordinates with membrane trafficking during cytokinesis, remains poorly understood. Here we show that Myo1 is a two-headed myosin II in Saccharomyces cerevisiae, and that Myo1 localizes to the division site via two distinct targeting signals in its tail that act sequentially during the cell cycle. Before cytokinesis, Myo1 localization depends on the septin-binding protein Bni5. During cytokinesis, Myo1 localization depends on the IQGAP Iqg1. We also show that the Myo1 tail is sufficient for promoting the assembly of a “headless” AMR, which guides membrane deposition and extracellular matrix remodeling at the division site. Our study establishes a biphasic targeting mechanism for myosin II and highlights an underappreciated role of the AMR in cytokinesis beyond force generation.

scholarly journals Distinct roles of Rho1, Cdc42, and Cyk3 in septum formation and abscission during yeast cytokinesis

2013 ◽  
Vol 202 (2) ◽  
pp. 311-329 ◽  
Author(s):  
Masayuki Onishi ◽  
Nolan Ko ◽  
Ryuichi Nishihama ◽  
John R. Pringle
Keyword(s):  
Small Gtpase ◽  
Dominant Negative ◽  
Cleavage Furrow ◽  
Genetic Screens ◽  
General Role ◽  
Guanosine Diphosphate ◽  
Septum Formation ◽  
Primary Septum ◽  
Ring Constriction ◽  
Guanosine Triphosphatase

In yeast and animal cytokinesis, the small guanosine triphosphatase (GTPase) Rho1/RhoA has an established role in formation of the contractile actomyosin ring, but its role, if any, during cleavage-furrow ingression and abscission is poorly understood. Through genetic screens in yeast, we found that either activation of Rho1 or inactivation of another small GTPase, Cdc42, promoted secondary septum (SS) formation, which appeared to be responsible for abscission. Consistent with this hypothesis, a dominant-negative Rho1 inhibited SS formation but not cleavage-furrow ingression or the concomitant actomyosin ring constriction. Moreover, Rho1 is temporarily inactivated during cleavage-furrow ingression; this inactivation requires the protein Cyk3, which binds Rho1-guanosine diphosphate via its catalytically inactive transglutaminase-like domain. Thus, unlike the active transglutaminases that activate RhoA, the multidomain protein Cyk3 appears to inhibit activation of Rho1 (and thus SS formation), while simultaneously promoting cleavage-furrow ingression through primary septum formation. This work suggests a general role for the catalytically inactive transglutaminases of fungi and animals, some of which have previously been implicated in cytokinesis.

scholarly journals Cleavage-furrow formation without F-actin in Chlamydomonas

2019 ◽  
Author(s):  
Masayuki Onishi ◽  
James G. Umen ◽  
Frederick R. Cross ◽  
John R. Pringle
Keyword(s):  
Cell Division ◽  
Chlamydomonas Reinhardtii ◽  
Green Alga ◽  
Eukaryotic Cells ◽  
Cleavage Furrow ◽  
Type Ii ◽  
Type Ii Myosin ◽  
In Cells

AbstractIt is widely believed that cleavage-furrow formation during cell division is driven by the contraction of a ring containing F-actin and type-II myosin. However, even in cells that have such rings, they are not always essential for furrow formation. Moreover, many taxonomically diverse eukaryotic cells divide by furrowing but have no type-II myosin, making it unlikely that an actomyosin ring drives furrowing. To explore this issue further, we have used one such organism, the green alga Chlamydomonas reinhardtii. We found that although F-actin is concentrated in the furrow region, none of the three myosins (of types VIII and XI) is localized there. Moreover, when F-actin was eliminated through a combination of a mutation and a drug, furrows still formed and the cells divided, although somewhat less efficiently than normal. Unexpectedly, division of the large Chlamydomonas chloroplast was delayed in the cells lacking F-actin; as this organelle lies directly in the path of the cleavage furrow, this delay may explain, at least in part, the delay in cell division itself. Earlier studies had shown an association of microtubules with the cleavage furrow, and we used a fluorescently tagged EB1 protein to show that at least the microtubule plus-ends are still associated with the furrows in the absence of F-actin, consistent with the possibility that the microtubules are important for furrow formation. We suggest that the actomyosin ring evolved as one way to improve the efficiency of a core process for furrow formation that was already present in ancestral eukaryotes.

Fission yeast Rng3p: an UCS-domain protein that mediates myosin II assembly during cytokinesis

2000 ◽  
Vol 113 (13) ◽  
pp. 2421-2432 ◽  
Author(s):  
K.C. Wong ◽  
N.I. Naqvi ◽  
Y. Iino ◽  
M. Yamamoto ◽  
M.K. Balasubramanian
Keyword(s):  
Fission Yeast ◽  
Fluorescent Protein ◽  
Type Ii ◽  
Division Site ◽  
Ring Assembly ◽  
Medial Cortex ◽  
Late Step ◽  
Green Fluorescent ◽  
Type Ii Myosin

Cell division in many eukaryotes, including the fission yeast Schizosaccharomyces pombe, utilizes a contractile actomyosin ring. In S. pombe, the actomyosin ring is assembled at the medial cortex upon entry into mitosis and constricts at the end of anaphase to guide the centripetal deposition of the septum. Despite identification of several structural components essential for actomyosin ring assembly, the interdependencies between these gene-products in the process of ring assembly are unknown. This study investigates the role of Rng3p, a member of the UCS-domain containing protein family (Unc-45p, Cro1p, She4p), in actomyosin ring assembly. Null mutants in rng3 resemble deletion mutants in the type II myosin heavy chain (myo2) and rng3(ts) mutants show strong negative interactions with the myo2-E1 mutant, suggesting that Rng3p is involved in modulating aspects of type II myosin function. Interestingly, a green fluorescent protein (GFP) tagged Rng3p fusion is detected at the division site in the myo2-E1 mutant, but not in other myo2-alleles, wild-type cells or in 18 other cytokinesis mutants. Assembly and maintenance of Rng3p at the division site in the myo2-E1 mutant requires F-actin. Rng3p is also required for the proper assembly of Myo2p and F-actin into a functional actomyosin ring but is not necessary for their accumulation at the division site. We conclude that Rng3p is a novel component of the F-actin cytoskeleton essential for a late step in actomyosin ring assembly and that it might monitor some aspect of type II myosin assembly during actomyosin ring construction.

scholarly journals Identification and functional analysis of the essential and regulatory light chains of the only type II myosin Myo1p in Saccharomyces cerevisiae

2004 ◽  
Vol 165 (6) ◽  
pp. 843-855 ◽  
Author(s):  
Jianying Luo ◽  
Elizabeth A. Vallen ◽  
Christopher Dravis ◽  
Serguei E. Tcheperegine ◽  
Becky Drees ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Point Mutation ◽  
Light Chain ◽  
Type Ii ◽  
Actin Ring ◽  
Major Function ◽  
Regulatory Light Chains ◽  
Essential Light Chain ◽  
Type V ◽  
Type Ii Myosin

Cytokinesis in Saccharomyces cerevisiae involves coordination between actomyosin ring contraction and septum formation and/or targeted membrane deposition. We show that Mlc1p, a light chain for Myo2p (type V myosin) and Iqg1p (IQGAP), is the essential light chain for Myo1p, the only type II myosin in S. cerevisiae. However, disruption or reduction of Mlc1p–Myo1p interaction by deleting the Mlc1p binding site on Myo1p or by a point mutation in MLC1, mlc1-93, did not cause any obvious defect in cytokinesis. In contrast, a different point mutation, mlc1-11, displayed defects in cytokinesis and in interactions with Myo2p and Iqg1p. These data suggest that the major function of the Mlc1p–Myo1p interaction is not to regulate Myo1p activity but that Mlc1p may interact with Myo1p, Iqg1p, and Myo2p to coordinate actin ring formation and targeted membrane deposition during cytokinesis. We also identify Mlc2p as the regulatory light chain for Myo1p and demonstrate its role in Myo1p ring disassembly, a function likely conserved among eukaryotes.

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