Mutants of the Drosophila ncd microtubule motor protein cause centrosomal and spindle pole defects in mitosis

Nonclaret disjunctional (ncd) is a kinesin-related microtubule motor protein required for meiotic and early mitotic chromosome distribution in Drosophila. ncd translocates on microtubules with the opposite polarity to kinesin, toward microtubule minus ends, and is associated with spindles in chromosome/spindle preparations. Here we report a new mutant of ncd caused by partial deletion of the predicted coiled-coil central stalk. The mutant protein exhibits a velocity of translocation and ability to generate torque in motility assays comparable to near full-length ncd, but only partially rescues a null mutant for chromosome mis-segregation. Antibody staining experiments show that the partial loss-of-function and null mutants cause centrosomal and spindle pole defects, including centrosome splitting and loss of centrosomes from spindle poles, and localize ncd to centrosomes as well as spindles of wild-type embryos. Association of ncd with spindles and centrosomes is microtubule- and cell cycle-dependent: inhibition of microtubule assembly with colchicine abolishes ncd staining and centrosomal staining is observed in prometaphase, metaphase and anaphase, but diminishes in late anaphase/telophase. The cell cycle dependence of centrosomal staining and the defects of mutants provide clear evidence for activity of the ncd motor protein near or at the spindle poles in mitosis. The ncd motor may interact with centrosomal microtubules and spindle fibers to attach centrosomes to spindle poles, and mediate poleward translocation (flux) of kinetochore fibers, a process that may underlie poleward movement of chromosomes in mitosis. Together with previous work, our findings indicate that ncd is important in maintaining spindle poles in mitosis as well as in meiosis.

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Mutants of the microtubule motor protein, nonclaret disjunctional, affect spindle structure and chromosome movement in meiosis and mitosis

Journal of Cell Science ◽

10.1242/jcs.101.3.547 ◽

1992 ◽

Vol 101 (3) ◽

pp. 547-559 ◽

Cited By ~ 3

Author(s):

M. Hatsumi ◽

S.A. Endow

Keyword(s):

Motor Protein ◽

Spindle Motor ◽

Meiotic Spindle ◽

Chromosome Movement ◽

Chromosome Distribution ◽

Early Embryos ◽

Microtubule Motor ◽

Microtubule Motor Protein ◽

Spindle Structure ◽

Meiosis I

The Drosophila microtubule motor protein, nonclaret disjunctional (ncd), is required for proper chromosome distribution in meiosis and mitosis. We have examined the meiotic and mitotic divisions in wild-type Drosophila oocytes and early embryos, and the effects of three ncd mutants (cand, ncd and ncdD) on spindle structure and chromosome movement. The ncd mutants cause abnormalities in spindle structure early in meiosis I, and abnormal chromosome configurations throughout meiosis I and II. Defective divisions continue in early embryos of the motor null mutant, cand, with abnormal early mitotic spindles. The effects of mutants on spindle structure suggest that ncd is required for proper meiotic spindle assembly, and may play a role in forming or maintaining spindle poles in meiosis. The disruption of normal meiotic and mitotic chromosome distribution by ncd mutants can be attributed to its role as a spindle motor, although a role for ncd as a chromosome-associated motor protein is not excluded. The ncd motor protein functions not only in meiosis, but also performs an active role in the early mitotic divisions of the embryo.

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The cell cycle state defines TACC3 as a regulator gene in glioblastoma

10.1101/2020.10.20.346643 ◽

2020 ◽

Author(s):

Holly Briggs ◽

Euan S. Polson ◽

Bronwyn K. Irving ◽

Alexandre Zougman ◽

Ryan K. Mathew ◽

...

Keyword(s):

Cell Cycle ◽

Gene Networks ◽

Coiled Coil ◽

Cell Lineage ◽

Progression Free Survival ◽

Low Grade ◽

Loss Of Function ◽

Feature Maps ◽

Clinical Databases ◽

Lineage Specificity

AbstractOverexpression and mitosis-promoting roles of Transforming acidic coiled-coil containing protein 3 (TACC3) are well-established in many cancers, including glioblastoma (GBM). However, the effector gene networks downstream of TACC3 remain poorly defined, partly due to an incomplete understanding of TACC3 cell lineage specificity and its dynamic role during the cell cycle. Here, we use a patient-derived GBM model to report that TACC3 predominantly resides in the GBM cell cytoplasm, while engaging in gene regulation temporally as defined by the cell cycle state. TACC3 loss-of-function, cell cycle stage-specific transcriptomics, and unsupervised self-organizing feature maps revealed pathways (including Hedgehog signalling) and individual genes (including HOTAIR) that exhibited anticorrelated expression phenotypes across interphase and mitosis. Furthermore, this approach identified a set of 22 TACC3-dependent transcripts in publicly-available clinical databases that predicted poor overall and progression-free survival in 162 GBM and 514 low-grade glioma patient samples. These findings uncover TACC3-dependent genes as a function of TACC3 cell cycle oscillation, which is important for TACC3-targeting strategies, and for predicting poor outcomes in brain cancer patients.

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The role of gamma-tubulin in mitotic spindle formation and cell cycle progression in Aspergillus nidulans

Journal of Cell Science ◽

10.1242/jcs.110.5.623 ◽

1997 ◽

Vol 110 (5) ◽

pp. 623-633 ◽

Cited By ~ 3

Author(s):

M.A. Martin ◽

S.A. Osmani ◽

B.R. Oakley

Keyword(s):

Cell Cycle ◽

Mitotic Spindle ◽

Cell Cycle Progression ◽

Spindle Pole ◽

Microtubule Assembly ◽

Cytoplasmic Microtubule ◽

Cycle Progression ◽

Spindle Microtubules ◽

Gamma Tubulin

gamma-Tubulin has been hypothesized to be essential for the nucleation of the assembly of mitotic spindle microtubules, but some recent results suggest that this may not be the case. To clarify the role of gamma-tubulin in microtubule assembly and cell-cycle progression, we have developed a novel variation of the gene disruption/heterokaryon rescue technique of Aspergillus nidulans. We have used temperature-sensitive cell-cycle mutations to synchronize germlings carrying a gamma-tubulin disruption and observe the phenotypes caused by the disruption in the first cell cycle after germination. Our results indicate that gamma-tubulin is absolutely required for the assembly of mitotic spindle microtubules, a finding that supports the hypothesis that gamma-tubulin is involved in spindle microtubule nucleation. In the absence of functional gamma-tubulin, nuclei are blocked with condensed chromosomes for about the length of one cell cycle before chromatin decondenses without nuclear division. Our results indicate that gamma-tubulin is not essential for progression from G1 to G2, for entry into mitosis nor for spindle pole body replication. It is also not required for reactivity of spindle pole bodies with the MPM-2 antibody which recognizes a phosphoepitope important to mitotic spindle formation. Finally, it does not appear to be absolutely required for cytoplasmic microtubule assembly but may play a role in the formation of normal cytoplasmic microtubule arrays.

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The organization of Golgi in Drosophila bristles requires microtubule motor protein function and a properly organized microtubule array

PLoS ONE ◽

10.1371/journal.pone.0223174 ◽

2019 ◽

Vol 14 (10) ◽

pp. e0223174

Author(s):

Anna Melkov ◽

Raju Baskar ◽

Rotem Shachal ◽

Yehonathan Alcalay ◽

Uri Abdu

Keyword(s):

Protein Function ◽

Motor Protein ◽

Microtubule Array ◽

Microtubule Motor ◽

Microtubule Motor Protein

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Use of GFP Fusions to a Microtubule Motor Protein to Analyze Spindle Dynamics in Live Oocytes and Embryos of Drosophila

Microscopy and Microanalysis ◽

10.1017/s1431927600007522 ◽

1997 ◽

Vol 3 (S2) ◽

pp. 127-128

Author(s):

S. A. Endow ◽

D. J. Komma

Keyword(s):

Fusion Proteins ◽

Motor Protein ◽

Spindle Motor ◽

Gene Fusions ◽

Mitotic Spindles ◽

Wild Type ◽

Early Embryos ◽

Microtubule Motor ◽

Microtubule Motor Protein ◽

Spindle Fibers

Ncd is a kinesin-related microtubule motor protein of Drosophila that plays essential roles in spindle assembly and function during meiosis in oocytes and mitosis in early embryos. Antibody staining experiments have localized the Ned motor protein to spindle fibers and spindle poles throughout the meiotic and early mitotic divisions, demonstrating that Ncd is a spindle motor.We have made ncd-gfp gene fusions with wild-type and S65T gfp and expressed the chimaeric genes in Drosophila to target GFP to the spindle. Transgenic Drosophila carrying the ncd-gfp gene fusions in an ncd null mutant background are wild type with respect to chromosome segregation, indicating that the Ncd-GFP fusion proteins can replace the function of wild-type Ncd. The Ncd-GFP fusion proteins in transgenic Drosophila are expressed under the regulation of the native ncd promoter.Analysis of live Drosophila oocytes and early embryos shows green fluorescent spindles, demonstrating association of Ncd-GFP with meiotic and mitotic spindles. In mitotic spindles, Ncd-GFP localizes to centrosomes (Fig. 1a) and spindle fibers (Fig. 1b).

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Sld5 Ensures Centrosomal Resistance to Congression Forces by Preserving Centriolar Satellites

Molecular and Cellular Biology ◽

10.1128/mcb.00371-17 ◽

2017 ◽

Vol 38 (2) ◽

Cited By ~ 1

Author(s):

Manpreet Kaur ◽

Raksha Devi ◽

Tanushree Ghosh ◽

Md Muntaz Khan ◽

Praveen Kumar ◽

...

Keyword(s):

Dna Replication ◽

Unknown Function ◽

Motor Protein ◽

Spindle Pole ◽

Replication Protein ◽

Traction Forces ◽

Centrosomal Protein ◽

Spindle Poles ◽

Chromosome Congression ◽

Pulling Forces

ABSTRACT The migration of chromosomes during mitosis is mediated primarily by kinesins that bind to the chromosomes and move along the microtubules, exerting pulling and pushing forces on the centrosomes. We report that a DNA replication protein, Sld5, localizes to the centrosomes, resisting the microtubular pulling forces experienced during chromosome congression. In the absence of Sld5, centriolar satellites, which normally cluster around the centrosomes, are dissipated throughout the cytoplasm, resulting in the loss of their known function of recruiting the centrosomal protein, pericentrin. We observed that Sld5-deficient centrosomes lacking pericentrin were unable to endure the CENP-E- and Kid-mediated microtubular forces that converge on the centrosomes during chromosome congression, resulting in monocentriolar and acentriolar spindle poles. The minus-end-directed kinesin-14 motor protein, HSET, sustains the traction forces that mediate centrosomal fragmentation in Sld5-depleted cells. Thus, we report that a DNA replication protein has an as yet unknown function of ensuring spindle pole resistance to traction forces exerted during chromosome congression.

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