TEDs Site Phosphorylation Regulates Myosin I Motor Activity And Function In Fission Yeast

ABSTRACTIn both unicellular and multicellular organisms, long-tailed class I myosins function in clathrin-mediated endocytosis. Myosin 1e (Myo1e) in vertebrates and Myo1 in fission yeast have similar domain organization, yet whether these proteins or their individual protein domains are functionally interchangeable remains unknown. In an effort to assess functional conservation of class I myosins, we tested whether human Myo1e could replace Myo1 in fission yeast Schizosaccharomyces pombe and found that it was unable to substitute for yeast Myo1. To determine if any individual protein domain is responsible for the inability of Myo1e to function in yeast, we created human-yeast myosin-I chimeras. By functionally testing these chimeric myosins in vivo, we concluded that the Myo1e motor domain is unable to function in yeast, even when combined with the yeast Myo1 tail and a full complement of yeast regulatory light chains. Conversely, the Myo1e tail, when attached to the yeast Myo1 motor domain, supports localization to actin patches and partially rescues the endocytosis defect in myo1Δ cells. Further dissection showed that both the TH1 and TH2-SH3 domains in the human Myo1e tail are required for localization and function of chimeric myosin-I at endocytic sites. Overall, this study provides insights into the role of individual myosin-I domains, expands the utility of fission yeast as a simple model system to study the effects of disease-associated MYO1E mutations, and supports a model of co-evolution between a myosin motor and its actin track.

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Roles of the novel coiled-coil protein Rng10 in septum formation during fission yeast cytokinesis

Molecular Biology of the Cell ◽

10.1091/mbc.e16-03-0156 ◽

2016 ◽

Vol 27 (16) ◽

pp. 2528-2541 ◽

Cited By ~ 6

Author(s):

Yajun Liu ◽

I-Ju Lee ◽

Mingzhai Sun ◽

Casey A. Lower ◽

Kurt W. Runge ◽

...

Keyword(s):

Fission Yeast ◽

Rho Gtpases ◽

Cellular Localization ◽

Affinity Purification ◽

Coiled Coil ◽

The Novel ◽

Septum Formation ◽

Division Site ◽

And Function

Rho GAPs are important regulators of Rho GTPases, which are involved in various steps of cytokinesis and other processes. However, regulation of Rho-GAP cellular localization and function is not fully understood. Here we report the characterization of a novel coiled-coil protein Rng10 and its relationship with the Rho-GAP Rga7 in fission yeast. Both rng10Δ and rga7Δ result in defective septum and cell lysis during cytokinesis. Rng10 and Rga7 colocalize on the plasma membrane at the cell tips during interphase and at the division site during cell division. Rng10 physically interacts with Rga7 in affinity purification and coimmunoprecipitation. Of interest, Rga7 localization is nearly abolished without Rng10. Moreover, Rng10 and Rga7 work together to regulate the accumulation and dynamics of glucan synthases for successful septum formation in cytokinesis. Our results show that cellular localization and function of the Rho-GAP Rga7 are regulated by a novel protein, Rng10, during cytokinesis in fission yeast.

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Big Lessons from Little Yeast: Budding and Fission Yeast Centrosome Structure, Duplication, and Function

Annual Review of Genetics ◽

10.1146/annurev-genet-120116-024733 ◽

2017 ◽

Vol 51 (1) ◽

pp. 361-383 ◽

Cited By ~ 34

Author(s):

Ann M. Cavanaugh ◽

Sue L. Jaspersen

Keyword(s):

Fission Yeast ◽

And Function

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The Putative RNA-Binding Protein Nrp1 Promotes the Loading of Kinesin-14/Klp2 to the Mitotic Spindle and Is Sequestered Into Heat-Induced Protein Aggregates in Fission Yeast

10.20944/preprints202103.0452.v1 ◽

2021 ◽

Author(s):

Masashi Yukawa ◽

Mitsuki Ohishi ◽

Yusuke Yamada ◽

Takashi Toda

Keyword(s):

Fission Yeast ◽

Rna Binding ◽

Binding Protein ◽

Rna Binding Protein ◽

Protein Aggregates ◽

Associated Proteins ◽

Spindle Microtubules ◽

The Stability ◽

And Function ◽

Post Transcriptional Regulation

Cells form a bipolar spindle during mitosis to ensure accurate chromosome segregation. Proper spindle architecture is established by a set of kinesin motors and microtubule-associated proteins. In most eukaryotes, kinesin-5 motors are essential for this process, and genetic or chemical inhibition of their activity leads to the emergence of monopolar spindles and cell death. However, these deficiencies can be rescued by simultaneous inactivation of kinesin-14 motors, as they counteract kinesin-5. We conducted detailed genetic analyses in fission yeast to understand the mechanisms driving spindle assembly in the absence of kinesin-5. Here we show that deletion of the nrp1 gene, which encodes a putative RNA-binding protein with unknown function, can rescue temperature sensitivity caused by cut7-22, a fission yeast kinesin-5 mutant. Interestingly, kinesin-14/Klp2 levels on the spindles in the cut7 mutants were significantly reduced by the nrp1 deletion, although the total levels of Klp2 and the stability of spindle microtubules remained unaffected. Moreover, RNA-binding motifs of Nrp1 are essential for its cytoplasmic localization and function. We have also found that a portion of Nrp1 is spatially and functionally sequestered by chaperone-based protein aggregates upon mild heat stress and limits cell division at high temperatures. We propose that Nrp1 might be involved in post-transcriptional regulation through its RNA-binding ability to promote the loading of Klp2 on the spindle microtubules.

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Threonine-11, Phosphorylated by Rad3 and ATM In Vitro, Is Required for Activation of Fission Yeast Checkpoint Kinase Cds1

Molecular and Cellular Biology ◽

10.1128/mcb.21.10.3398-3404.2001 ◽

2001 ◽

Vol 21 (10) ◽

pp. 3398-3404 ◽

Cited By ~ 36

Author(s):

Katsunori Tanaka ◽

Michael N. Boddy ◽

Xiao-Bo Chen ◽

Clare H. McGowan ◽

Paul Russell

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Fission Yeast ◽

Ataxia Telangiectasia ◽

Null Mutant ◽

Ataxia Telangiectasia Mutated ◽

Tolerance Mechanisms ◽

Checkpoint Kinase ◽

And Function

ABSTRACT Fission yeast Cds1 is phosphorylated and activated when DNA replication is interrupted by nucleotide starvation or DNA damage. Cds1 enforces the S-M checkpoint that couples mitosis (M) to the completion of DNA synthesis (S). Cds1 also controls replicational stress tolerance mechanisms. Cds1 is regulated by a group of proteins that includes Rad3, a kinase related to human checkpoint kinase ATM (ataxia telangiectasia mutated). ATM phosphorylates serine or threonine followed by glutamine (SQ or TQ). Here we show that in vitro, Rad3 and ATM phosphorylate the N-terminal domain of Cds1 at the motif T11Q12. Substitution of threonine-11 with alanine (T11A) abolished Cds1 activation that occurs when DNA replication is inhibited by hydroxyurea (HU) treatment. Thecds1-T11A mutant was profoundly sensitive to HU, although not quite as sensitive as a cds1− null mutant. Cds1T11A was unable to enforce the S-M checkpoint. These results strongly suggest that Rad3-dependent phosphorylation of Cds1 at threonine-11 is required for Cds1 activation and function.

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Actin and nuclear myosin I are associated with RNAP II and function in gene transcription

Chinese Science Bulletin ◽

10.1007/s11434-007-0126-z ◽

2007 ◽

Vol 52 (6) ◽

pp. 766-770 ◽

Cited By ~ 1

Author(s):

XiaoJuan Zhu ◽

BaiQu Huang ◽

XingZhi Wang ◽

Shui Hao ◽

XianLu Zeng

Keyword(s):

Gene Transcription ◽

Myosin I ◽

Nuclear Myosin I ◽

Rnap Ii ◽

And Function

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Cells with Broken Left–Right Symmetry: Roles of Intrinsic Cell Chirality in Left–Right Asymmetric Epithelial Morphogenesis

Symmetry ◽

10.3390/sym11040505 ◽

2019 ◽

Vol 11 (4) ◽

pp. 505 ◽

Cited By ~ 4

Author(s):

Sosuke Utsunomiya ◽

So Sakamura ◽

Takeshi Sasamura ◽

Tomoki Ishibashi ◽

Chinami Maeda ◽

...

Keyword(s):

Animal Species ◽

Animal Cell ◽

Fruit Fly ◽

Cellular Level ◽

Structure And Function ◽

Epithelial Morphogenesis ◽

Asymmetric Structure ◽

Molecular Chirality ◽

Myosin I ◽

And Function

Chirality is a fundamental feature in biology, from the molecular to the organismal level. An animal has chirality in the left–right asymmetric structure and function of its body. In general, chirality occurring at the molecular and organ/organism scales has been studied separately. However, recently, chirality was found at the cellular level in various species. This “cell chirality” can serve as a link between molecular chirality and that of an organ or animal. Cell chirality is observed in the structure, motility, and cytoplasmic dynamics of cells and the mechanisms of cell chirality formation are beginning to be understood. In all cases studied so far, proteins that interact chirally with F-actin, such as formin and myosin I, play essential roles in cell chirality formation or the switching of a cell’s enantiomorphic state. Thus, the chirality of F-actin may represent the ultimate origin of cell chirality. Links between cell chirality and left–right body asymmetry are also starting to be revealed in various animal species. In this review, the mechanisms of cell chirality formation and its roles in left–right asymmetric development are discussed, with a focus on the fruit fly Drosophila, in which many of the pioneering studies were conducted.

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Roles of putative Rho-GEF Gef2 in division-site positioning and contractile-ring function in fission yeast cytokinesis

Molecular Biology of the Cell ◽

10.1091/mbc.e11-09-0800 ◽

2012 ◽

Vol 23 (7) ◽

pp. 1181-1195 ◽

Cited By ~ 36

Author(s):

Yanfang Ye ◽

I-Ju Lee ◽

Kurt W. Runge ◽

Jian-Qiu Wu

Keyword(s):

Fission Yeast ◽

Rho Gtpases ◽

Nucleotide Exchange ◽

Contractile Ring ◽

Division Plane ◽

Cell Functions ◽

Division Site ◽

Ring Assembly ◽

And Function ◽

Rho Gef

Cytokinesis is crucial for integrating genome inheritance and cell functions. In multicellular organisms, Rho-guanine nucleotide exchange factors (GEFs) and Rho GTPases are key regulators of division-plane specification and contractile-ring formation during cytokinesis, but how they regulate early steps of cytokinesis in fission yeast remains largely unknown. Here we show that putative Rho-GEF Gef2 and Polo kinase Plo1 coordinate to control the medial cortical localization and function of anillin-related protein Mid1. The division-site positioning defects of gef2∆ plo1-ts18 double mutant can be partially rescued by increasing Mid1 levels. We find that Gef2 physically interacts with the Mid1 N-terminus and modulates Mid1 cortical binding. Gef2 localization to cortical nodes and the contractile ring depends on its last 145 residues, and the DBL-homology domain is important for its function in cytokinesis. Our data suggest the interaction between Rho-GEFs and anillins is an important step in the signaling pathways during cytokinesis. In addition, Gef2 also regulates contractile-ring function late in cytokinesis and may negatively regulate the septation initiation network. Collectively, we propose that Gef2 facilitates and stabilizes Mid1 binding to the medial cortex, where the localized Mid1 specifies the division site and induces contractile-ring assembly.

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Deciphering the 3D Structure and Function of Phosphofructokinase from Fission Yeast

Microscopy and Microanalysis ◽

10.1017/s143192761400796x ◽

2014 ◽

Vol 20 (S3) ◽

pp. 1246-1247

Author(s):

Shaun Benjamin ◽

Michael Radermacher ◽

Teresa Ruiz

Keyword(s):

Fission Yeast ◽

3D Structure ◽

Structure And Function ◽

And Function

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

Molecular Biology of the Cell ◽

10.1091/mbc.e09-10-0852 ◽

2010 ◽

Vol 21 (6) ◽

pp. 989-1000 ◽

Cited By ~ 55

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.

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