Excess histone H3 is a competitive Chk1 inhibitor that controls cell-cycle remodeling in the early Drosophila embryo

The Concentration of Nuf, a Rab11 Effector, at the Microtubule-organizing Center Is Cell Cycle–regulated, Dynein-dependent, and Coincides with Furrow Formation

Molecular Biology of the Cell ◽

10.1091/mbc.e07-02-0146 ◽

2007 ◽

Vol 18 (9) ◽

pp. 3313-3322 ◽

Cited By ~ 36

Author(s):

Blake Riggs ◽

Barbara Fasulo ◽

Anne Royou ◽

Sarah Mische ◽

Jian Cao ◽

...

Keyword(s):

Cell Cycle ◽

Small Gtpase ◽

Drosophila Embryo ◽

Microtubule Organizing Center ◽

Recycling Endosome ◽

Actin Organization ◽

Protein Levels ◽

Nuclear Fallout ◽

Early Drosophila Embryo ◽

Organizing Center

Animal cytokinesis relies on membrane addition as well as acto-myosin–based constriction. Recycling endosome (RE)-derived vesicles are a key source of this membrane. Rab11, a small GTPase associated with the RE and involved in vesicle targeting, is required for elongation of the cytokinetic furrow. In the early Drosophila embryo, Nuclear-fallout (Nuf), a Rab11 effector, promotes vesicle-mediated membrane delivery and actin organization at the invaginating furrow. Although Rab11 maintains a relatively constant localization at the microtubule-organizing center (MTOC), Nuf is present at the MTOC only during the phases of the cell cycle in which furrow invagination occurs. We demonstrate that Nuf protein levels remain relatively constant throughout the cell cycle, suggesting that Nuf is undergoing cycles of concentration and dispersion from the MTOC. Microtubules, but not microfilaments, are required for proper MTOC localization of Nuf and Rab11. The MTOC localization of Nuf also relies on Dynein. Immunoprecipitation experiments demonstrate that Nuf and Dynein physically interact. In accord with these findings, and in contrast to previous reports, we demonstrate that microtubules are required for proper metaphase furrow formation. We propose that the cell cycle–regulated, Dynein-dependent recruitment of Nuf to the MTOC influences the timing of RE-based vesicle delivery to the invaginating furrows.

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The cell cycle-dependent localization of the CP190 centrosomal protein is determined by the coordinate action of two separable domains.

The Journal of Cell Biology ◽

10.1083/jcb.131.5.1261 ◽

1995 ◽

Vol 131 (5) ◽

pp. 1261-1273 ◽

Cited By ~ 40

Author(s):

K Oegema ◽

W G Whitfield ◽

B Alberts

Keyword(s):

Cell Cycle ◽

Amino Acids ◽

Nuclear Localization ◽

Fusion Proteins ◽

Drosophila Embryo ◽

Dependent Manner ◽

Fluorescence Confocal Microscopy ◽

Early Drosophila Embryo ◽

A Cell ◽

Cycle Dependent

CP190, a protein of 1,096 amino acids from Drosophila melanogaster, oscillates in a cell cycle-specific manner between the nucleus during interphase, and the centrosome during mitosis. To characterize the regions of CP190 responsible for its dynamic behavior, we injected rhodamine-labeled fusion proteins spanning most of CP190 into early Drosophila embryos, where their localizations were characterized using time-lapse fluorescence confocal microscopy. A single bipartite 19-amino acid nuclear localization signal was detected that causes nuclear localization. Robust centrosomal localization is conferred by a separate region of 124 amino acids; two adjacent, nonoverlapping fusion proteins containing distinct portions of this region show weaker centrosomal localization. Fusion proteins that contain both nuclear and centrosomal localization sequences oscillate between the nucleus and the centrosome in a manner identical to native CP190. Fusion proteins containing only the centrosome localization sequence are found at centrosomes throughout the cell cycle, suggesting that CP190 is actively recruited away from the centrosome by its movement into the nucleus during interphase. Both native and bacterially expressed CP190 cosediment with microtubules in vitro. Tests with fusion proteins show that the domain responsible for microtubule binding overlaps the domain required for centrosomal localization. CP60, a protein identified by its association with CP190, also localizes to centrosomes and to nuclei in a cell cycle-dependent manner. Experiments in which colchicine is used to depolymerize microtubules in the early Drosophila embryo demonstrate that both CP190 and CP60 are able to attain and maintain their centrosomal localization in the absence of microtubules.

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Influence of cyclin type and dose on mitotic entry and progression in the early Drosophila embryo

The Journal of Cell Biology ◽

10.1083/jcb.200810012 ◽

2009 ◽

Vol 184 (5) ◽

pp. 639-646 ◽

Cited By ~ 25

Author(s):

Mark L. McCleland ◽

Jeffrey A. Farrell ◽

Patrick H. O'Farrell

Keyword(s):

Cell Cycle ◽

Drosophila Melanogaster ◽

Drosophila Embryo ◽

Cell Cycle Regulators ◽

Mitotic Progression ◽

Mitotic Entry ◽

Early Drosophila Embryo ◽

Real Time Visualization ◽

The One

Cyclins are key cell cycle regulators, yet few analyses test their role in timing the events that they regulate. We used RNA interference and real-time visualization in embryos to define the events regulated by each of the three mitotic cyclins of Drosophila melanogaster, CycA, CycB, and CycB3. Each individual and pairwise knockdown results in distinct mitotic phenotypes. For example, mitosis without metaphase occurs upon knockdown of CycA and CycB. To separate the role of cyclin levels from the influences of cyclin type, we knocked down two cyclins and reduced the gene dose of the one remaining cyclin. This reduction did not prolong interphase but instead interrupted mitotic progression. Mitotic prophase chromosomes formed, centrosomes divided, and nuclei exited mitosis without executing later events. This prompt but curtailed mitosis shows that accumulation of cyclin function does not directly time mitotic entry in these early embryonic cycles and that cyclin function can be sufficient for some mitotic events although inadequate for others.

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Chk1 and Wee1 kinases coordinate DNA replication, chromosome condensation, and anaphase entry

Molecular Biology of the Cell ◽

10.1091/mbc.e11-10-0832 ◽

2012 ◽

Vol 23 (6) ◽

pp. 1047-1057 ◽

Cited By ~ 23

Author(s):

Barbara Fasulo ◽

Carol Koyama ◽

Kristina R. Yu ◽

Ellen M. Homola ◽

Tao S. Hsieh ◽

...

Keyword(s):

Cell Cycle ◽

Dna Replication ◽

Anticancer Agents ◽

Spindle Assembly Checkpoint ◽

Metaphase Plate ◽

Chromosome Condensation ◽

Drosophila Embryo ◽

X Rays ◽

Checkpoint Activation ◽

Early Drosophila Embryo

Defects in DNA replication and chromosome condensation are common phenotypes in cancer cells. A link between replication and condensation has been established, but little is known about the role of checkpoints in monitoring chromosome condensation. We investigate this function by live analysis, using the rapid division cycles in the early Drosophila embryo. We find that S-phase and topoisomerase inhibitors delay both the initiation and the rate of chromosome condensation. These cell cycle delays are mediated by the cell cycle kinases chk1 and wee1. Inhibitors that cause severe defects in chromosome condensation and congression on the metaphase plate result in delayed anaphase entry. These delays are mediated by wee1 and are not the result of spindle assembly checkpoint activation. In addition, we provide the first detailed live analysis of the direct effect of widely used anticancer agents (aclarubicin, ICRF-193, VM26, doxorubicin, camptothecin, aphidicolin, hydroxyurea, cisplatin, mechlorethamine and x-rays) on key nuclear and cytoplasmic cell cycle events.

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