The Drosophila ncd microtubule motor protein is spindle-associated in meiotic and mitotic cells

The nonclaret disjunctional (ncd) protein is required for normal chromosome distribution in oocytes and early embryos. Mutants of ncd cause frequent nondisjunction and loss of chromosomes, suggesting a role for the protein in spindle function or chromosome movement in meiosis and early mitosis. The ncd protein contains a region of predicted sequence similarity to the microtubule motor protein, kinesin. In vitro motility assays have demonstrated that ncd is a motor that unexpectedly moves toward the minus ends of microtubules, opposite to the direction of kinesin movement. Using antibodies directed against nonconserved regions of the protein, we have localized the ncd motor protein to the meiotic and early mitotic spindle, and to spindles in a mitotically dividing cultured cell line. Its presence in the spindle of meiotic and mitotic cells implies a role for the protein as a spindle motor. The motor may play an essential role in establishing spindle bipolarity in meiosis.

Download Full-text

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.

Download Full-text

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).

Download Full-text

Chemical subdomains within the kinetochore domain of isolated CHO mitotic chromosomes.

The Journal of Cell Biology ◽

10.1083/jcb.114.2.285 ◽

1991 ◽

Vol 114 (2) ◽

pp. 285-294 ◽

Cited By ~ 58

Author(s):

L Wordeman ◽

E R Steuer ◽

M P Sheetz ◽

T Mitchison

Keyword(s):

Indirect Immunofluorescence ◽

Motor Protein ◽

Chromosome Translocation ◽

Mitotic Chromosomes ◽

Cell Biol ◽

Microtubule Motor ◽

Specific Antigens ◽

Microtubule Motor Protein

We have used indirect immunofluorescence in combination with correlative EM to subdivide the mammalian kinetochore into two domains based on the localization of specific antigens. We demonstrate here that the fibrous corona on the distal face of the kinetochore plate contains tubulin (previously shown by Mitchison, T. J., and M. W. Kirschner. 1985. J. Cell Biol. 101:755-765) and the minus end-directed, ATP-dependent microtubule motor protein, dynein; whereas a 50-kD CREST antigen is located internal to these components in the kinetochore. Tubulin and dynein can be extracted from the kinetochore by 150 mM KI, leaving other, as yet uncharacterized, components of the kinetochore corona intact. Microtubules and tubulin subunits will associate with kinetochores in vitro after extraction with 150 mM KI, suggesting that other functionally significant, corona-associated molecules remain unextracted. Our results suggest that the corona region of the kinetochore contains the machinery for chromosome translocation along microtubules.

Download Full-text

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

Download Full-text

Mitochondrial Association of a Plus End–Directed Microtubule Motor Expressed during Mitosis in Drosophila

The Journal of Cell Biology ◽

10.1083/jcb.136.5.1081 ◽

1997 ◽

Vol 136 (5) ◽

pp. 1081-1090 ◽

Cited By ~ 88

Author(s):

Andrea J. Pereira ◽

Brian Dalby ◽

Russell J. Stewart ◽

Stephen J. Doxsey ◽

Lawrence S.B. Goldstein

Keyword(s):

System Development ◽

Sequence Similarity ◽

Cell Types ◽

Nervous System Development ◽

In Vitro Motility Assay ◽

Specific Expression ◽

In Vitro Motility ◽

Amino Terminal ◽

Specific Expression Pattern

The kinesin superfamily is a large group of proteins (kinesin-like proteins [KLPs]) that share sequence similarity with the microtubule (MT) motor kinesin. Several members of this superfamily have been implicated in various stages of mitosis and meiosis. Here we report our studies on KLP67A of Drosophila. DNA sequence analysis of KLP67A predicts an MT motor protein with an amino-terminal motor domain. To prove this directly, KLP67A expressed in Escherichia coli was shown in an in vitro motility assay to move MTs in the plus direction. We also report expression analyses at both the mRNA and protein level, which implicate KLP67A in the localization of mitochondria in undifferentiated cell types. In situ hybridization studies of the KLP67A mRNA during embryogenesis and larval central nervous system development indicate a proliferation-specific expression pattern. Furthermore, when affinity-purified anti-KLP67A antisera are used to stain blastoderm embryos, mitochondria in the region of the spindle asters are labeled. These data suggest that KLP67A is a mitotic motor of Drosophila that may have the unique role of positioning mitochondria near the spindle.

Download Full-text

Microtubule flux in mitosis is independent of chromosomes, centrosomes, and antiparallel microtubules.

Molecular Biology of the Cell ◽

10.1091/mbc.5.2.217 ◽

1994 ◽

Vol 5 (2) ◽

pp. 217-226 ◽

Cited By ~ 37

Author(s):

K E Sawin ◽

T J Mitchison

Keyword(s):

Basic Element ◽

Low Light ◽

Self Organized ◽

Egg Extracts ◽

Microtubule Motor ◽

Xenopus Egg ◽

Xenopus Egg Extracts ◽

Microtubule Motor Protein ◽

Poleward Flux

We investigated the mechanism of poleward microtubule flux in the mitotic spindle by generating spindle subassemblies in Xenopus egg extracts in vitro and assaying their ability to flux by photoactivation of fluorescence and low-light multichannel fluorescence video-microscopy. We find that monopolar intermediates of in vitro spindle assembly (half-spindles) exhibit normal poleward flux, as do astral microtubule arrays induced by the addition of dimethyl sulfoxide to egg extracts in the absence of both chromosomes and conventional centrosomes. Immunodepletion of the kinesin-related microtubule motor protein Eg5, a candidate flux motor, suggests that Eg5 is not required for flux. These results suggest that poleward flux is a basic element of microtubule behavior exhibited by even simple self-organized microtubule arrays and presumably underlies the most elementary levels of spindle morphogenesis.

Download Full-text