CEP110 and ninein are located in a specific domain of the centrosome associated with centrosome maturation

2002 ◽  
Vol 115 (9) ◽  
pp. 1825-1835 ◽  
Author(s):  
Young Y. Ou ◽  
Gary J. Mack ◽  
Meifeng Zhang ◽  
Jerome B. Rattner
Keyword(s):  
Cell Cycle ◽  
Protein Interactions ◽  
Protein Function ◽  
Similar Distribution ◽  
Microtubule Organizing Center ◽  
Specific Domain ◽  
The Reformation ◽  
Mother And Daughter ◽  
Organizing Center ◽  
Pericentriolar Material

The mammalian centrosome consists of a pair of centrioles surrounded by pericentriolar material (PCM). The architecture and composition of the centrosome, especially the PCM, changes during the cell cycle. Recently, a subset of PCM proteins have been shown to be arranged in a tubular conformation with an open and a closed end within the centrosome. The presence of such a specific configuration can be used as a landmark for mapping proteins in both a spatial and a temporal fashion. Such mapping studies can provide information about centrosome organization, protein dynamics,protein-protein interactions as well as protein function. In this study, the centrosomal proteins CEP110 and ninein were mapped in relationship to the tubular configuration. Both proteins were found to exhibit a similar distribution pattern. In the mother centrosome, they were found at both ends of the centrosome tube, including the site of centrosome duplication. However,in the daughter centrosome they were present only at the closed end. At the closed end of the mother and daughter centrosome tube, both CEP110 and ninein co-localized with the centriolar protein CEP250/c-Nap1, which confirms ninein's centriole association and places CEP110 in association with this structure. Importantly, the appearance of CEP110 and ninein at the open end of the daughter centrosome occurred during the telophase-G1 transition of the next cell cycle, concomitant with the maturation of the daughter centrosome into a mother centrosome. Microinjection of antibodies against either CEP110 or ninein into metaphase HeLa cells disrupted the reformation of the tubular conformation of proteins within the centrosome following cell division and consequently led to dispersal of centrosomal material throughout the cytosol. Further, microinjection of antibodies to either CEP110 or ninein into metaphase PtK2 cells not only disrupted the tubular configuration within the centrosome but also affected the centrosome's ability to function as a microtubule organizing center (MTOC). This MTOC function was also disrupted when the antibodies were injected into postmitotic cells. Taken together, our results indicate that: (1) a population of CEP110 and ninein is located in a specific domain within the centrosome, which corresponds to the open end of the centrosome tube and is the site of protein addition associated with maturation of a daughter centrosome into a mother centrosome; and (2) the addition of CEP110 and ninein are essential for the reformation of specific aspects of the interphase centrosome architecture following mitosis as well as being required for the centrosome to function as a MTOC.

scholarly journals A two-step mechanism for the inactivation of microtubule organizing center function at the centrosome

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Jérémy Magescas ◽  
Jenny C Zonka ◽  
Jessica L Feldman
Keyword(s):  
Cell Cycle ◽  
Mitotic Spindle ◽  
Outer Sphere ◽  
Microtubule Organizing Center ◽  
C Elegans ◽  
Mitotic Kinases ◽  
Pulling Forces ◽  
Organizing Center ◽  
Pericentriolar Material ◽  
Step Mechanism

The centrosome acts as a microtubule organizing center (MTOC), orchestrating microtubules into the mitotic spindle through its pericentriolar material (PCM). This activity is biphasic, cycling through assembly and disassembly during the cell cycle. Although hyperactive centrosomal MTOC activity is a hallmark of some cancers, little is known about how the centrosome is inactivated as an MTOC. Analysis of endogenous PCM proteins in C. elegans revealed that the PCM is composed of partially overlapping territories organized into an inner and outer sphere that are removed from the centrosome at different rates and using different behaviors. We found that phosphatases oppose the addition of PCM by mitotic kinases, ultimately catalyzing the dissolution of inner sphere PCM proteins at the end of mitosis. The nature of the PCM appears to change such that the remaining aging PCM outer sphere is mechanically ruptured by cortical pulling forces, ultimately inactivating MTOC function at the centrosome.

scholarly journals Breaking the ties that bind: New advances in centrosome biology

2012 ◽  
Vol 197 (1) ◽  
pp. 11-18 ◽  
Author(s):  
Balca R. Mardin ◽  
Elmar Schiebel
Keyword(s):  
Cell Cycle ◽  
Cancer Cells ◽  
Recent Work ◽  
Genomic Instability ◽  
Centrosome Duplication ◽  
Animal Cells ◽  
Microtubule Organizing Center ◽  
Centrosome Separation ◽  
Organizing Center ◽  
Pericentriolar Material

The centrosome, which consists of two centrioles and the surrounding pericentriolar material, is the primary microtubule-organizing center (MTOC) in animal cells. Like chromosomes, centrosomes duplicate once per cell cycle and defects that lead to abnormalities in the number of centrosomes result in genomic instability, a hallmark of most cancer cells. Increasing evidence suggests that the separation of the two centrioles (disengagement) is required for centrosome duplication. After centriole disengagement, a proteinaceous linker is established that still connects the two centrioles. In G2, this linker is resolved (centrosome separation), thereby allowing the centrosomes to separate and form the poles of the bipolar spindle. Recent work has identified new players that regulate these two processes and revealed unexpected mechanisms controlling the centrosome cycle.

scholarly journals The centrosomin CM2 domain is a multi-functional binding domain with distinct cell cycle roles

2017 ◽  
Author(s):  
Y. Rose Citron ◽  
Carey J. Fagerstrom ◽  
Bettina Keszthelyi ◽  
Bo Huang ◽  
Nasser M Rusan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Interactions ◽  
Intramolecular Interaction ◽  
Sequence Divergence ◽  
Nucleation Site ◽  
Binding Assays ◽  
Microtubule Organizing Center ◽  
Organizing Center ◽  
Cycle Dependent

AbstractThe centrosome serves as the main microtubule-organizing center in metazoan cells, yet despite its functional importance, little is known mechanistically about the structure and organizational principles that dictate protein organization in the centrosome. In particular, the protein-protein interactions that allow for the massive structural transition between the tightly organized interphase centrosome and the highly expanded matrix-like arrangement of the mitotic centrosome have been largely uncharacterized. Among the proteins that undergo a major transition is the Drosophila melanogaster protein centrosomin that contains a conserved carboxyl terminus motif, CM2. Recent crystal structures have shown this motif to be dimeric and capable of forming an intramolecular interaction with a central region of centrosomin. Here we use a combination of in-cell microscopy and in vitro oligomer assessment to show that dimerization is not necessary for CM2 recruitment to the centrosome and that CM2 alone undergoes a significant cell cycle dependent rearrangement. We use NMR binding assays to confirm this intramolecular interaction and show that residues involved in solution interactions are consistent with the published crystal structure and identify L1137 as critical for binding. Additionally, we show for the first time an in vitro interaction of CM2 with the Drosophila pericentrin-like-protein that exploits the same set of residues as the intramolecular interaction. Furthermore, NMR experiments reveal a calcium sensitive interaction between CM2 and calmodulin. Although unexpected because of sequence divergence, this suggests that centrosomin-mediated assemblies, like the mammalian pericentrin, may be calcium regulated. From these results we suggest a model where during interphase CM2 interacts with pericentrin-like-protein to form a layer of centrosomin around the centriole wall and that at the onset of mitosis this population acts as a nucleation site of intramolecular centrosomin interactions that support the expansion into the metaphase matrix.

scholarly journals A two-step mechanism for the inactivation of microtubule organizing center function at the centrosome

2018 ◽  
Author(s):  
Jérémy Magescas ◽  
Jennifer C. Zonka ◽  
Jessica L. Feldman
Keyword(s):  
Cell Cycle ◽  
Mitotic Spindle ◽  
Microtubule Organizing Center ◽  
Step Process ◽  
C Elegans ◽  
Pulling Forces ◽  
Organizing Center ◽  
Pericentriolar Material ◽  
And Behavior ◽  
Step Mechanism

SummaryDuring mitosis, the centrosome acts as a microtubule organizing center (MTOC), orchestrating microtubules into the mitotic spindle through its pericentriolar material (PCM). This activity is biphasic, cycling through assembly and disassembly during the cell cycle. Although hyperactive centrosomal MTOC activity is a hallmark of some cancers, little is known about how the centrosome is inactivated as an MTOC. Analysis of endogenous PCM proteins in C. elegans revealed that the PCM is composed of distinct protein territories that are removed from the centrosome at different rates and using different behaviors. Inhibition of PP2A phosphatases stabilized the PCM and perturbation of cortical pulling forces altered the timing and behavior by which proteins were removed from the centrosome. These data indicate that PCM disassembly is a two-step process, beginning with a phosphatase-dependent dissolution of PCM proteins followed by the ejection of ruptured PCM by cortical forces, ultimately inactivating MTOC function at the centrosome.

scholarly journals Daughter Centriole Elongation Is Controlled by Proteolysis

2010 ◽  
Vol 21 (22) ◽  
pp. 3942-3951 ◽  
Author(s):  
Nina Korzeniewski ◽  
Rolando Cuevas ◽  
Anette Duensing ◽  
Stefan Duensing
Keyword(s):  
Mammalian Cells ◽  
Gene Products ◽  
Microtubule Organizing Center ◽  
Future Studies ◽  
Microtubule Stability ◽  
Organizing Center ◽  
Pericentriolar Material ◽  
Substrate Cleavage ◽  
Interfering Rna ◽  
Daughter Centriole

The centrosome is the major microtubule-organizing center of most mammalian cells and consists of a pair of centrioles embedded in pericentriolar material. Before mitosis, the two centrioles duplicate and two new daughter centrioles form adjacent to each preexisting maternal centriole. After initiation of daughter centriole synthesis, the procentrioles elongate in a process that is poorly understood. Here, we show that inhibition of cellular proteolysis by Z-L3VS or MG132 induces abnormal elongation of daughter centrioles to approximately 4 times their normal length. This activity of Z-L3VS or MG132 was found to correlate with inhibition of intracellular protease-mediated substrate cleavage. Using a small interfering RNA screen, we identified a total of nine gene products that either attenuated (seven) or promoted (two) abnormal Z-L3VS–induced daughter centriole elongation. Our hits included known regulators of centriole length, including CPAP and CP110, but, interestingly, several proteins involved in microtubule stability and anchoring as well as centrosome cohesion. This suggests that nonproteasomal functions, specifically inhibition of cellular proteases, may play an important and underappreciated role in the regulation of centriole elongation. They also highlight the complexity of daughter centriole length control and provide a framework for future studies to dissect the molecular details of this process.

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

2007 ◽  
Vol 18 (9) ◽  
pp. 3313-3322 ◽  
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.

A protein related to brain microtubule-associated protein MAP1B is a component of the mammalian centrosome

1994 ◽  
Vol 107 (2) ◽  
pp. 601-611 ◽  
Author(s):  
J.E. Dominguez ◽  
B. Buendia ◽  
C. Lopez-Otin ◽  
C. Antony ◽  
E. Karsenti ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Cultured Cells ◽  
Polyclonal Antibodies ◽  
Microtubule Organizing Center ◽  
Microtubule Associated Proteins ◽  
Organizing Center ◽  
Associated Proteins ◽  
Pericentriolar Material ◽  
Peptide Maps

The centrosome is the main microtubule organizing center of mammalian cells. Structurally, it is composed of a pair of centrioles surrounded by a fibro-granular material (the pericentriolar material) from which microtubules are nucleated. However, the nature of centrosomal molecules involved in microtubules nucleation is still obscure. Since brain microtubule-associated proteins (MAPs) lower the critical tubulin concentration required for microtubule nucleation in tubulin solution in vitro, we have examined their possible association with centrosomes. By immunofluorescence, monoclonal and polyclonal antibodies raised against MAP1B stain the centrosome in cultured cells as well as purified centrosomes, whereas antibodies raised against MAP2 give a completely negative reaction. The MAP1B-related antigen is localized to the pericentriolar material as revealed by immunoelectron microscopy. In preparations of purified centrosomes analyzed on poly-acrylamide gels, a protein that migrates as brain MAP1B is present. After blotting on nitrocellulose, it is decorated by anti-MAP1B antibodies and the amino acid sequence of proteolytic fragments of this protein is similar to brain MAP1B. Moreover, brain MAP1B and its centrosomal counterpart share the same phosphorylation features and have similar peptide maps. These data strongly suggest that a protein homologue to MAP1B is present in centrosomes and it is a good candidate for being involved in the nucleating activity of the pericentriolar material.

scholarly journals Kinetics and intermediates of marginal band reformation: evidence for peripheral determinants of microtubule organization.

1984 ◽  
Vol 99 (1) ◽  
pp. 70s-75s ◽  
Author(s):  
M Miller ◽  
F Solomon
Keyword(s):  
Cell Types ◽  
Microtubule Assembly ◽  
Cell Periphery ◽  
Microtubule Organizing Center ◽  
Microtubule Organization ◽  
Mature Erythrocyte ◽  
The Reformation ◽  
Marginal Band ◽  
Organizing Center

The microtubules of the mature erythrocyte of the chicken are confined to a band at the periphery. Whole-mount electron microscopy after extraction reveals that the number of microtubules in each cell is almost the same. All the microtubules can be depolymerized by incubation in the cold, and the marginal band can be quantitatively and qualitatively reformed by return to 39 degrees C. These properties allow the reformation of the marginal band to be treated as an in vivo microtubule assembly reaction. The kinetics of this reaction and the intermediates detected during reformation suggest a mechanism of microtubule organization that is distinct from that observed in other cell types. Apparently only one or two growing microtubule ends are available for assembly--assembly is only detected at the cell periphery, even at early times--and there is no evidence of the participation of a microtubule-organizing center.

scholarly journals The centriolar satellite protein Cfap53/Ccdc11 facilitates the formation of the first zygotic microtubule organizing center in the zebrafish embryo

2020 ◽  
Author(s):  
Sven Willekers ◽  
Federico Tessadori ◽  
Babet van der Vaart ◽  
Heiko Henning ◽  
Riccardo Stucchi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Danio Rerio ◽  
Mitotic Spindle ◽  
Sperm Cell ◽  
Zebrafish Embryo ◽  
Animal Species ◽  
Microtubule Organizing Center ◽  
Microtubule Network ◽  
Cell Divisions ◽  
Organizing Center

AbstractIn embryos from most animal species a zygotic centrosome is assembled by the centriole derived from the sperm cell and pericentriolar proteins present in the oocyte. This zygotic centrosome acts as a microtubule organizing center (MTOC) to assemble the mitotic spindle in the first and all subsequent cell divisions. As MTOC formation has been studied mainly in adult cells, very little is known about the formation of the first zygotic MTOC. Here we find that zebrafish (Danio rerio) embryos lacking maternal or paternal Cfap53, a centriolar satellite protein, arrest during the first cell cycle due to a failure in proper formation of the mitotic spindle. During the first cell cycle Cfap53 co-localizes with γ-tubulin and other centrosomal and centriolar satellite proteins to the very large MTOC. Furthermore, we find that γ-tubulin localization to the MTOC is impaired in the absence of Cfap53 or when the microtubule network is disrupted. Based on these results we propose a model in which maternal and paternal Cfap53 participates in the organization of the first zygotic MTOC of the embryo. Once the zygotic MTOC is formed, Cfap53 is dispensable for MTOC formation and integrity in subsequent cell divisions.

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