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

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

Journal of Cell Science ◽

10.1242/jcs.103.4.1013 ◽

1992 ◽

Vol 103 (4) ◽

pp. 1013-1020 ◽

Cited By ~ 15

Author(s):

M. Hatsumi ◽

S.A. Endow

Keyword(s):

Sequence Similarity ◽

Motor Protein ◽

Spindle Motor ◽

In Vitro Motility ◽

Early Embryos ◽

Microtubule Motor ◽

Spindle Function ◽

Microtubule Motor Protein ◽

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.

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Spindle Dynamics during Meiosis in Drosophila Oocytes

The Journal of Cell Biology ◽

10.1083/jcb.137.6.1321 ◽

1997 ◽

Vol 137 (6) ◽

pp. 1321-1336 ◽

Cited By ~ 85

Author(s):

Sharyn A. Endow ◽

Donald J. Komma

Keyword(s):

Fluorescent Protein ◽

Meiotic Spindle ◽

Oocyte Activation ◽

New Information ◽

Microtubule Motor ◽

Meiotic Spindles ◽

Green Fluorescent ◽

Microtubule Motor Protein ◽

Meiosis I

Mature oocytes of Drosophila are arrested in metaphase of meiosis I. Upon activation by ovulation or fertilization, oocytes undergo a series of rapid changes that have not been directly visualized previously. We report here the use of the Nonclaret disjunctional (Ncd) microtubule motor protein fused to the green fluorescent protein (GFP) to monitor changes in the meiotic spindle of live oocytes after activation in vitro. Meiotic spindles of metaphase-arrested oocytes are relatively stable, however, meiotic spindles of in vitro–activated oocytes are highly dynamic: the spindles elongate, rotate around their long axis, and undergo an acute pivoting movement to reorient perpendicular to the oocyte surface. Many oocytes spontaneously complete the meiotic divisions, permitting visualization of progression from meiosis I to II. The movements of the spindle after oocyte activation provide new information about the dynamic changes in the spindle that occur upon re-entry into meiosis and completion of the meiotic divisions. Spindles in live oocytes mutant for a lossof-function ncd allele fused to gfp were also imaged. The genesis of spindle defects in the live mutant oocytes provides new insights into the mechanism of Ncd function in the spindle during the meiotic divisions.

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