scholarly journals Regulation of EDEN-dependent deadenylation of Aurora A/Eg2-derived mRNA via phosphorylation and dephosphorylation in Xenopus laevis egg extracts

2003 ◽  
Vol 116 (13) ◽  
pp. 2697-2705 ◽  
Author(s):  
L. Detivaud
Keyword(s):  
Xenopus Laevis ◽  
Aurora A ◽  
Egg Extracts

scholarly journals Aurora A phosphorylation of TACC3/maskin is required for centrosome-dependent microtubule assembly in mitosis

2005 ◽  
Vol 170 (7) ◽  
pp. 1047-1055 ◽  
Author(s):  
Kazuhisa Kinoshita ◽  
Tim L. Noetzel ◽  
Laurence Pelletier ◽  
Karl Mechtler ◽  
David N. Drechsel ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Xenopus Laevis ◽  
Microtubule Assembly ◽  
Aurora A ◽  
Specific Mechanism ◽  
Microtubule Polymerization ◽  
Egg Extracts ◽  
Microtubule Growth

Centrosomes act as sites of microtubule growth, but little is known about how the number and stability of microtubules emanating from a centrosome are controlled during the cell cycle. We studied the role of the TACC3–XMAP215 complex in this process by using purified proteins and Xenopus laevis egg extracts. We show that TACC3 forms a one-to-one complex with and enhances the microtubule-stabilizing activity of XMAP215 in vitro. TACC3 enhances the number of microtubules emanating from mitotic centrosomes, and its targeting to centrosomes is regulated by Aurora A–dependent phosphorylation. We propose that Aurora A regulation of TACC3 activity defines a centrosome-specific mechanism for regulation of microtubule polymerization in mitosis.

scholarly journals Quantitative NMR analysis of the protein G B1 domain in Xenopus laevis egg extracts and intact oocytes

2006 ◽  
Vol 103 (32) ◽  
pp. 11904-11909 ◽  
Author(s):  
P. Selenko ◽  
Z. Serber ◽  
B. Gadea ◽  
J. Ruderman ◽  
G. Wagner
Keyword(s):  
Xenopus Laevis ◽  
Protein G ◽  
Nmr Analysis ◽  
Quantitative Nmr ◽  
B1 Domain ◽  
Egg Extracts

scholarly journals Characterization of the TPX2 Domains Involved in Microtubule Nucleation and Spindle Assembly in Xenopus Egg Extracts

2004 ◽  
Vol 15 (12) ◽  
pp. 5318-5328 ◽  
Author(s):  
Stéphane Brunet ◽  
Teresa Sardon ◽  
Timo Zimmerman ◽  
Torsten Wittmann ◽  
Rainer Pepperkok ◽  
...  
Keyword(s):  
Spindle Assembly ◽  
Aurora A Kinase ◽  
Aurora A ◽  
Microtubule Nucleation ◽  
Terminal Domain ◽  
Large N ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Domain Functional ◽  
Xenopus Egg Extracts

TPX2 has multiple functions during mitosis, including microtubule nucleation around the chromosomes and the targeting of Xklp2 and Aurora A to the spindle. We have performed a detailed domain functional analysis of TPX2 and found that a large N-terminal domain containing the Aurora A binding peptide interacts directly with and nucleates microtubules in pure tubulin solutions. However, it cannot substitute the endogenous TPX2 to support microtubule nucleation in response to Ran guanosine triphosphate (GTP) and spindle assembly in egg extracts. By contrast, a large C-terminal domain of TPX2 that does not bind directly to pure microtubules and does not bind Aurora A kinase rescues microtubule nucleation in response to RanGTP and spindle assembly in TPX2-depleted extract. These and previous results suggest that under physiological conditions, TPX2 is essential for microtubule nucleation around chromatin and functions in a network of other molecules, some of which also are regulated by RanGTP.

scholarly journals Bimodal Interaction of Mammalian Polo-Like Kinase 1 and a Centrosomal Scaffold, Cep192, in the Regulation of Bipolar Spindle Formation

2015 ◽  
Vol 35 (15) ◽  
pp. 2626-2640 ◽  
Author(s):  
Lingjun Meng ◽  
Jung-Eun Park ◽  
Tae-Sung Kim ◽  
Eun Hye Lee ◽  
Suk-Youl Park ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Spindle Assembly ◽  
Human Cells ◽  
Mitotic Progression ◽  
Spindle Formation ◽  
Content Type ◽  
Bipolar Spindle ◽  
Egg Extracts ◽  
Cultured Human Cells ◽  
Bipolar Spindles

Serving as microtubule-organizing centers, centrosomes play a key role in forming bipolar spindles. The mechanism of how centrosomes promote bipolar spindle assembly in various organisms remains largely unknown. A recent study withXenopus laevisegg extracts suggested that the Plk1 ortholog Plx1 interacts with the phospho-T46 (p-T46) motif ofXenopusCep192 (xCep192) to form an xCep192-mediated xAurA-Plx1 cascade that is critical for bipolar spindle formation. Here, we demonstrated that in cultured human cells, Cep192 recruits AurA and Plk1 in a cooperative manner, and this event is important for the reciprocal activation of AurA and Plk1. Strikingly, Plk1 interacted with Cep192 through either the p-T44 (analogous toXenopusp-T46) or the newly identified p-S995 motif via its C-terminal noncatalytic polo-box domain. The interaction between Plk1 and the p-T44 motif was prevalent in the presence of Cep192-bound AurA, whereas the interaction of Plk1 with the p-T995 motif was preferred in the absence of AurA binding. Notably, the loss of p-T44- and p-S995-dependent Cep192-Plk1 interactions induced an additive defect in recruiting Plk1 and γ-tubulin to centrosomes, which ultimately led to a failure in proper bipolar spindle formation and mitotic progression. Thus, we propose that Plk1 promotes centrosome-based bipolar spindle formation by forming two functionally nonredundant complexes with Cep192.

The NIMA-related kinase X-Nek2B is required for efficient assembly of the zygotic centrosome in Xenopus laevis

2000 ◽  
Vol 113 (11) ◽  
pp. 1973-1984 ◽  
Author(s):  
A.M. Fry ◽  
P. Descombes ◽  
C. Twomey ◽  
R. Bacchieri ◽  
E.A. Nigg
Keyword(s):  
Cell Cycle ◽  
Xenopus Laevis ◽  
Basal Body ◽  
Chromatin Condensation ◽  
Embryonic Cells ◽  
Serine Threonine Kinase ◽  
Cell Cycles ◽  
Functional Studies ◽  
Mammalian Cell Cycle ◽  
Egg Extracts

Nek2 is a mammalian cell cycle-regulated serine/threonine kinase that belongs to the family of proteins related to NIMA of Aspergillus nidulans. Functional studies in diverse species have implicated NIMA-related kinases in G(2)/M progression, chromatin condensation and centrosome regulation. To directly address the requirements for vertebrate Nek2 kinases in these cell cycle processes, we have turned to the biochemically-tractable system provided by Xenopus laevis egg extracts. Following isolation of a Xenopus homologue of Nek2, called X-Nek2B, we found that X-Nek2B abundance and activity remained constant through the first mitotic cycle implying a fundamental difference in Nek2 regulation between embryonic and somatic cell cycles. Removal of X-Nek2B from extracts did not disturb either entry into mitosis or the accompanying condensation of chromosomes providing no support for a requirement for Nek2 in these processes at least in embryonic cells. In contrast, X-Nek2B localized to centrosomes of adult Xenopus cells and was rapidly recruited to the basal body of Xenopus sperm following incubation in egg extracts. Recruitment led to phosphorylation of the X-Nek2B kinase. Most importantly, depletion of X-Nek2B from extracts significantly delayed both the assembly of microtubule asters and the recruitment of gamma-tubulin to the basal body. Hence, these studies demonstrate that X-Nek2B is required for efficient assembly of a functional zygotic centrosome and highlight the possibility of multiple roles for vertebrate Nek2 kinases in the centrosome cycle.

scholarly journals Histone H1 is essential for mitotic chromosome architecture and segregation in Xenopus laevis egg extracts

2005 ◽  
Vol 169 (6) ◽  
pp. 859-869 ◽  
Author(s):  
Thomas J. Maresca ◽  
Benjamin S. Freedman ◽  
Rebecca Heald
Keyword(s):  
Xenopus Laevis ◽  
Metaphase Plate ◽  
Histone H1 ◽  
Linker Histone ◽  
Chromosome Architecture ◽  
Linker Histone H1 ◽  
Egg Extracts ◽  
Chromatin Proteins ◽  
And Function

During cell division, condensation and resolution of chromosome arms and the assembly of a functional kinetochore at the centromere of each sister chromatid are essential steps for accurate segregation of the genome by the mitotic spindle, yet the contribution of individual chromatin proteins to these processes is poorly understood. We have investigated the role of embryonic linker histone H1 during mitosis in Xenopus laevis egg extracts. Immunodepletion of histone H1 caused the assembly of aberrant elongated chromosomes that extended off the metaphase plate and outside the perimeter of the spindle. Although functional kinetochores assembled, aligned, and exhibited poleward movement, long and tangled chromosome arms could not be segregated in anaphase. Histone H1 depletion did not significantly affect the recruitment of known structural or functional chromosomal components such as condensins or chromokinesins, suggesting that the loss of H1 affects chromosome architecture directly. Thus, our results indicate that linker histone H1 plays an important role in the structure and function of vertebrate chromosomes in mitosis.

scholarly journals Inhibition of protein synthesis in frog (Xenopus laevis) egg extracts by an antibody against phosphatidylinositol 4,5-bisphosphate

1996 ◽  
Vol 317 (3) ◽  
pp. 643-646
Author(s):  
Thomas J. KEATING ◽  
Kiyoko FUKAMI ◽  
Kenneth R. ROBINSON
Keyword(s):  
Protein Synthesis ◽  
Xenopus Laevis ◽  
Similar Reduction ◽  
Inhibition Of Protein Synthesis ◽  
Egg Extracts ◽  
Hydrolysis Of

The antibody kt10, which is directed against the phospholipid PtdIns(4,5)P2, inhibits protein synthesis when added to cytosolic extracts prepared from frog eggs. Addition of stable analogues of diacylglycerol and Ins(1,4,5)P3 failed to rescue the inhibition of translation, suggesting that the effect of the antibody was not to block hydrolysis of PtdIns(4,5)P2. Neomycin, which also binds PtdIns(4,5)P2, produced a similar reduction in protein-synthesis levels in the extract system, supporting the idea that it is the interaction of the antibody with PtdIns(4,5)P2 that is producing the effect.

scholarly journals Hepatoma Up-Regulated Protein Is Required for Chromatin-induced Microtubule Assembly Independently of TPX2

2008 ◽  
Vol 19 (11) ◽  
pp. 4900-4908 ◽  
Author(s):  
Claudia M. Casanova ◽  
Sofia Rybina ◽  
Hideki Yokoyama ◽  
Eric Karsenti ◽  
Iain W. Mattaj
Keyword(s):  
Xenopus Laevis ◽  
Detailed Analysis ◽  
Spindle Assembly ◽  
Microtubule Assembly ◽  
Spindle Microtubule ◽  
Microtubule Stabilization ◽  
Egg Extracts ◽  
Spindle Midzone ◽  
Microtubule Growth ◽  
Spindle Microtubules

The production of RanGTP around chromosomes is crucial for spindle microtubule assembly in mitosis. Previous work has shown that hepatoma up-regulated protein (HURP) is a Ran target, required for microtubule stabilization and spindle organization. Here we report a detailed analysis of HURP function in Xenopus laevis mitotic egg extracts. HURP depletion severely impairs bipolar spindle assembly around chromosomes: the few spindles that do form show a significant decrease in microtubule density at the spindle midzone. HURP depletion does not interfere with microtubule growth from purified centrosomes, but completely abolishes microtubule assembly induced by chromatin beads or RanGTP. Simultaneous depletion of the microtubule destabilizer MCAK with HURP does not rescue the phenotype, demonstrating that the effect of HURP is not to antagonize the destabilization activity of MCAK. Although the phenotype of HURP depletion closely resembles that reported for TPX2 depletion, we find no evidence that TPX2 and HURP physically interact or that they influence each other in their effects on spindle microtubules. Our data indicate that HURP and TPX2 have nonredundant functions essential for chromatin-induced microtubule assembly.

Repair of psoralen interstrand cross-links in Xenopus laevis egg extracts is highly mutagenic

2005 ◽  
Vol 336 (1) ◽  
pp. 69-75 ◽  
Author(s):  
Xiaoyan Lu ◽  
Nianxiang Zhang ◽  
Karen Vasquez ◽  
Michelle Barton ◽  
Randy Legerski
Keyword(s):  
Xenopus Laevis ◽  
Egg Extracts ◽  
Interstrand Cross Links ◽  
Cross Links
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