scholarly journals Fission yeast pkl1 is a kinesin-related protein involved in mitotic spindle function.

1996 ◽  
Vol 7 (10) ◽  
pp. 1639-1655 ◽  
Author(s):  
A L Pidoux ◽  
M LeDizet ◽  
W Z Cande
Keyword(s):  
Fission Yeast ◽  
Mitotic Spindle ◽  
Related Protein ◽  
Microtubule Organization ◽  
Spindle Poles ◽  
Similar Phenotype ◽  
Increased Sensitivity ◽  
Spindle Function ◽  
Related Proteins ◽  
Very High

We have used anti-peptide antibodies raised against highly conserved regions of the kinesin motor domain to identify kinesin-related proteins in the fission yeast Schizosaccharomyces pombe. Here we report the identification of a new kinesin-related protein, which we have named pkl1. Sequence homology and domain organization place pkl1 in the Kar3/ncd subfamily of kinesin-related proteins. Bacterially expressed pkl1 fusion proteins display microtubule-stimulated ATPase activity, nucleotide-sensitive binding, and bundling of microtubules. Immunofluorescence studies with affinity-purified antibodies indicate that the pkl1 protein localizes to the nucleus and the mitotic spindle. Pkl1 null mutants are viable but have increased sensitivity to microtubule-disrupting drugs. Disruption of pkl1+ suppresses mutations in another kinesin-related protein, cut7, which is known to act in the spindle. Overexpression of pkl1 to very high levels causes a similar phenotype to that seen in cut7 mutants: V-shaped and star-shaped microtubule structures are observed, which we interpret to be spindles with unseparated spindle poles. These observations suggest that pkl1 and cut7 provide opposing forces in the spindle. We propose that pkl1 functions as a microtubule-dependent motor that is involved in microtubule organization in the mitotic spindle.

scholarly journals Stu1p Is Physically Associated with β-Tubulin and Is Required for Structural Integrity of the Mitotic Spindle

2002 ◽  
Vol 13 (6) ◽  
pp. 1881-1892 ◽  
Author(s):  
Hongwei Yin ◽  
Liru You ◽  
Danielle Pasqualone ◽  
Kristen M. Kopski ◽  
Tim C. Huffaker
Keyword(s):  
Mitotic Spindle ◽  
Structural Integrity ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Yeast Saccharomyces Cerevisiae ◽  
Spindle Poles ◽  
Balance Of Forces ◽  
Related Proteins ◽  
Β Tubulin

Formation of the bipolar mitotic spindle relies on a balance of forces acting on the spindle poles. The primary outward force is generated by the kinesin-related proteins of the BimC family that cross-link antiparallel interpolar microtubules and slide them past each other. Here, we provide evidence that Stu1p is also required for the production of this outward force in the yeast Saccharomyces cerevisiae. In the temperature-sensitive stu1–5mutant, spindle pole separation is inhibited, and preanaphase spindles collapse, with their previously separated poles being drawn together. The temperature sensitivity of stu1–5 can be suppressed by doubling the dosage of Cin8p, a yeast BimC kinesin–related protein. Stu1p was observed to be a component of the mitotic spindle localizing to the midregion of anaphase spindles. It also binds to microtubules in vitro, and we have examined the nature of this interaction. We show that Stu1p interacts specifically with β-tubulin and identify the domains required for this interaction on both Stu1p and β-tubulin. Taken together, these findings suggest that Stu1p binds to interpolar microtubules of the mitotic spindle and plays an essential role in their ability to provide an outward force on the spindle poles.

scholarly journals Fission Yeast mto2p Regulates Microtubule Nucleation by the Centrosomin-related Protein mto1p

2005 ◽  
Vol 16 (6) ◽  
pp. 3040-3051 ◽  
Author(s):  
Itaru Samejima ◽  
Paula C. C. Lourenço ◽  
Hilary A. Snaith ◽  
Kenneth E. Sawin
Keyword(s):  
Fission Yeast ◽  
Insertional Mutagenesis ◽  
Spindle Pole ◽  
Related Protein ◽  
Microtubule Nucleation ◽  
Microtubule Organization ◽  
Cytoplasmic Microtubule ◽  
Division Site ◽  
Cell Extracts

From an insertional mutagenesis screen, we isolated a novel gene, mto2+, involved in microtubule organization in fission yeast. mto2Δ strains are viable but exhibit defects in interphase microtubule nucleation and in formation of the postanaphase microtubule array at the end of mitosis. The mto2Δ defects represent a subset of the defects displayed by cells deleted for mto1+ (also known as mod20+ and mbo1+), a centrosomin-related protein required to recruit the γ-tubulin complex to cytoplasmic microtubule-organizing centers (MTOCs). We show that mto2p colocalizes with mto1p at MTOCs throughout the cell cycle and that mto1p and mto2p coimmunoprecipitate from cytoplasmic extracts. In vitro studies suggest that mto2p binds directly to mto1p. In mto2Δ mutants, although some aspects of mto1p localization are perturbed, mto1p can still localize to spindle pole bodies and the cell division site and to “satellite” particles on interphase microtubules. In mto1Δ mutants, localization of mto2p to all of these MTOCs is strongly reduced or absent. We also find that in mto2Δ mutants, cytoplasmic forms of the γ-tubulin complex are mislocalized, and the γ-tubulin complex no longer coimmunoprecipitates with mto1p from cell extracts. These experiments establish mto2p as a major regulator of mto1p-mediated microtubule nucleation by the γ-tubulin complex.

scholarly journals A Novel Role of Dma1 in Regulating Forespore Membrane Assembly and Sporulation in Fission Yeast

2010 ◽  
Vol 21 (24) ◽  
pp. 4349-4360 ◽  
Author(s):  
Wen-zhu Li ◽  
Zhi-yong Yu ◽  
Peng-fei Ma ◽  
Yamei Wang ◽  
Quan-wen Jin
Keyword(s):  
Fission Yeast ◽  
Mitotic Spindle ◽  
Nuclear Division ◽  
Scaffold Proteins ◽  
Related Protein ◽  
Membrane Assembly ◽  
Meiotic Progression ◽  
Septation Initiation Network ◽  
Specific Proteins

In fission yeast Schizosaccharomyces pombe, a diploid mother cell differentiates into an ascus containing four haploid ascospores following meiotic nuclear divisions, through a process called sporulation. Several meiosis-specific proteins of fission yeast have been identified to play essential roles in meiotic progression and sporulation. We report here an unexpected function of mitotic spindle checkpoint protein Dma1 in proper spore formation. Consistent with its function in sporulation, expression of dma1+ is up-regulated during meiosis I and II. We showed that Dma1 localizes to the SPB during meiosis and the maintenance of this localization at meiosis II depends on septation initiation network (SIN) scaffold proteins Sid4 and Cdc11. Cells lacking Dma1 display defects associated with sporulation but not nuclear division, leading frequently to formation of asci with fewer spores. Our genetic analyses support the notion that Dma1 functions in parallel with the meiosis-specific Sid2-related protein kinase Slk1/Mug27 and the SIN signaling during sporulation, possibly through regulating proper forespore membrane assembly. Our studies therefore revealed a novel function of Dma1 in regulating sporulation in fission yeast.

scholarly journals Novel Roles for Saccharomyces cerevisiae Mitotic Spindle Motors

1999 ◽  
Vol 147 (2) ◽  
pp. 335-350 ◽  
Author(s):  
Frank R. Cottingham ◽  
Larisa Gheber ◽  
Dana L. Miller ◽  
M. Andrew Hoyt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Spindle ◽  
Structural Integrity ◽  
Cytoplasmic Dynein ◽  
Spindle Positioning ◽  
Bipolar Spindle ◽  
Accessory Factor ◽  
Spindle Function ◽  
Related Proteins ◽  
Spindle Structure

The single cytoplasmic dynein and five of the six kinesin-related proteins encoded by Saccharomyces cerevisiae participate in mitotic spindle function. Some of the motors operate within the nucleus to assemble and elongate the bipolar spindle. Others operate on the cytoplasmic microtubules to effect spindle and nuclear positioning within the cell. This study reveals that kinesin-related Kar3p and Kip3p are unique in that they perform roles both inside and outside the nucleus. Kar3p, like Kip3p, was found to be required for spindle positioning in the absence of dynein. The spindle positioning role of Kar3p is performed in concert with the Cik1p accessory factor, but not the homologous Vik1p. Kar3p and Kip3p were also found to overlap for a function essential for the structural integrity of the bipolar spindle. The cytoplasmic and nuclear roles of both these motors could be partially substituted for by the microtubule-destabilizing agent benomyl, suggesting that these motors perform an essential microtubule-destabilizing function. In addition, we found that yeast cell viability could be supported by as few as two microtubule-based motors: the BimC-type kinesin Cin8p, required for spindle structure, paired with either Kar3p or Kip3p, required for both spindle structure and positioning.

scholarly journals Mdb1, a Fission Yeast Homolog of Human MDC1, Modulates DNA Damage Response and Mitotic Spindle Function

PLoS ONE ◽  
2014 ◽  
Vol 9 (5) ◽  
pp. e97028 ◽  
Author(s):  
Yi Wei ◽  
Hai-Tao Wang ◽  
Yonggong Zhai ◽  
Paul Russell ◽  
Li-Lin Du
Keyword(s):  
Dna Damage ◽  
Fission Yeast ◽  
Dna Damage Response ◽  
Mitotic Spindle ◽  
Damage Response ◽  
Spindle Function ◽  
Yeast Homolog

scholarly journals Ase1p Organizes Antiparallel Microtubule Arrays during Interphase and Mitosis in Fission Yeast

2005 ◽  
Vol 16 (4) ◽  
pp. 1756-1768 ◽  
Author(s):  
Isabelle Loïodice ◽  
Jayme Staub ◽  
Thanuja Gangi Setty ◽  
Nam-Phuong T. Nguyen ◽  
Anne Paoletti ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Mitotic Spindle ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Microtubule Organization ◽  
Cellular Processes ◽  
Spindle Midzone ◽  
Fluorescence Live Cell Imaging

Proper microtubule organization is essential for cellular processes such as organelle positioning during interphase and spindle formation during mitosis. The fission yeast Schizosaccharomyces pombe presents a good model for understanding microtubule organization. We identify fission yeast ase1p, a member of the conserved ASE1/PRC1/MAP65 family of microtubule bundling proteins, which functions in organizing the spindle midzone during mitosis. Using fluorescence live cell imaging, we show that ase1p localizes to sites of microtubule overlaps associated with microtubule organizing centers at both interphase and mitosis. ase1Δ mutants fail to form overlapping antiparallel microtubule bundles, leading to interphase nuclear positioning defects, and premature mitotic spindle collapse. FRAP analysis revealed that interphase ase1p at overlapping microtubule minus ends is highly dynamic. In contrast, mitotic ase1p at microtubule plus ends at the spindle midzone is more stable. We propose that ase1p functions to organize microtubules into overlapping antiparallel bundles both in interphase and mitosis and that ase1p may be differentially regulated through the cell cycle.

scholarly journals ZYG-9, A Caenorhabditis elegans Protein Required for Microtubule Organization and Function, Is a Component of Meiotic and Mitotic Spindle Poles

1998 ◽  
Vol 141 (5) ◽  
pp. 1159-1168 ◽  
Author(s):  
Lisa R. Matthews ◽  
Philip Carter ◽  
Danielle Thierry-Mieg ◽  
Ken Kemphues
Keyword(s):  
Caenorhabditis Elegans ◽  
Mitotic Spindle ◽  
Meiotic Spindle ◽  
Microtubule Organization ◽  
Mitotic Apparatus ◽  
Meiotic Chromosomes ◽  
Spindle Poles ◽  
Common Activity ◽  
Early Anaphase ◽  
And Function

We describe the molecular characterization of zyg-9, a maternally acting gene essential for microtubule organization and function in early Caenorhabditis elegans embryos. Defects in zyg-9 mutants suggest that the zyg-9 product functions in the organization of the meiotic spindle and the formation of long microtubules. One-cell zyg-9 embryos exhibit both meiotic and mitotic spindle defects. Meiotic spindles are disorganized, pronuclear migration fails, and the mitotic apparatus forms at the posterior, orients incorrectly, and contains unusually short microtubules. We find that zyg-9 encodes a component of the meiotic and mitotic spindle poles. In addition to the strong staining of spindle poles, we consistently detect staining in the region of the kinetochore microtubules at metaphase and early anaphase in mitotic spindles. The ZYG-9 signal at the mitotic centrosomes is not reduced by nocodazole treatment, indicating that ZYG-9 localization to the mitotic centrosomes is not dependent upon long astral microtubules. Interestingly, in embryos lacking an organized meiotic spindle, produced either by nocodazole treatment or mutations in the mei-1 gene, ZYG-9 forms a halo around the meiotic chromosomes. The protein sequence shows partial similarity to a small set of proteins that also localize to spindle poles, suggesting a common activity of the proteins.

scholarly journals Myosin-10 and actin filaments are essential for mitotic spindle function

2008 ◽  
Vol 182 (1) ◽  
pp. 77-88 ◽  
Author(s):  
Sarah Woolner ◽  
Lori L. O'Brien ◽  
Christiane Wiese ◽  
William M. Bement
Keyword(s):  
Mitotic Spindle ◽  
Actin Filaments ◽  
Spindle Pole ◽  
Mitotic Spindles ◽  
Unconventional Myosin ◽  
Spindle Poles ◽  
Chromosome Partitioning ◽  
Meiotic Spindles ◽  
Spindle Function

Mitotic spindles are microtubule-based structures responsible for chromosome partitioning during cell division. Although the roles of microtubules and microtubule-based motors in mitotic spindles are well established, whether or not actin filaments (F-actin) and F-actin–based motors (myosins) are required components of mitotic spindles has long been controversial. Based on the demonstration that myosin-10 (Myo10) is important for assembly of meiotic spindles, we assessed the role of this unconventional myosin, as well as F-actin, in mitotic spindles. We find that Myo10 localizes to mitotic spindle poles and is essential for proper spindle anchoring, normal spindle length, spindle pole integrity, and progression through metaphase. Furthermore, we show that F-actin localizes to mitotic spindles in dynamic cables that surround the spindle and extend between the spindle and the cortex. Remarkably, although proper anchoring depends on both F-actin and Myo10, the requirement for Myo10 in spindle pole integrity is F-actin independent, whereas F-actin and Myo10 actually play antagonistic roles in maintenance of spindle length.

scholarly journals An Asp–CaM complex is required for centrosome–pole cohesion and centrosome inheritance in neural stem cells

2015 ◽  
Vol 211 (5) ◽  
pp. 987-998 ◽  
Author(s):  
Todd Schoborg ◽  
Allison L. Zajac ◽  
Carey J. Fagerstrom ◽  
Rodrigo X. Guillen ◽  
Nasser M. Rusan
Keyword(s):  
Stem Cells ◽  
Cell Polarity ◽  
Mitotic Spindle ◽  
Function Analysis ◽  
Tissue Homeostasis ◽  
Cell Cortex ◽  
Cross Link ◽  
Spindle Poles ◽  
Spindle Microtubules ◽  
Spindle Function

The interaction between centrosomes and mitotic spindle poles is important for efficient spindle formation, orientation, and cell polarity. However, our understanding of the dynamics of this relationship and implications for tissue homeostasis remains poorly understood. Here we report that Drosophila melanogaster calmodulin (CaM) regulates the ability of the microcephaly-associated protein, abnormal spindle (Asp), to cross-link spindle microtubules. Both proteins colocalize on spindles and move toward spindle poles, suggesting that they form a complex. Our binding and structure–function analysis support this hypothesis. Disruption of the Asp–CaM interaction alone leads to unfocused spindle poles and centrosome detachment. This behavior leads to randomly inherited centrosomes after neuroblast division. We further show that spindle polarity is maintained in neuroblasts despite centrosome detachment, with the poles remaining stably associated with the cell cortex. Finally, we provide evidence that CaM is required for Asp’s spindle function; however, it is completely dispensable for Asp’s role in microcephaly suppression.

scholarly journals Kinesin-Related Proteins at Mitotic Spindle Poles: Function and Regulation

Cell ◽  
1996 ◽  
Vol 85 (7) ◽  
pp. 943-946 ◽  
Author(s):  
Claire E Walczak ◽  
Timothy J Mitchison
Keyword(s):  
Mitotic Spindle ◽  
Spindle Poles ◽  
Related Proteins
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