Centriole-less pericentriolar material serves as a microtubule organizing center at the base of C. elegans sensory cilia

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.

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Centriole-less pericentriolar material serves as a microtubule organizing center at the base of C. elegans sensory cilia

10.1101/2020.08.20.260414 ◽

2020 ◽

Author(s):

Jérémy Magescas ◽

Sani Eskinazi ◽

Michael V. Tran ◽

Jessica L. Feldman

Keyword(s):

Sensory Neurons ◽

Super Resolution ◽

Spatial Separation ◽

Animal Cells ◽

Microtubule Organizing Center ◽

Tissue Specific ◽

C Elegans ◽

Organizing Center ◽

Pericentriolar Material ◽

Specific Degradation

SummaryDuring mitosis in animal cells, the centrosome acts as a microtubule organizing center (MTOC) to assemble the mitotic spindle. MTOC function at the centrosome is driven by proteins within the pericentriolar material (PCM), however the molecular complexity of the PCM makes it difficult to differentiate the proteins required for MTOC activity from other centrosomal functions. We used the natural spatial separation of PCM proteins during mitotic exit to identify a minimal module of proteins required for centrosomal MTOC function in C. elegans. Using tissue specific degradation, we show that SPD-5, the functional homolog of CDK5RAP2, is essential for embryonic mitosis while SPD-2/CEP192 and PCMD-1, which are essential in the zygote, are dispensable. Surprisingly, although the centriole is known to be degraded in the ciliated sensory neurons in C. elegans [1-3], we find evidence for “centriole-less PCM” at the base of cilia and use this structure as a minimal testbed to dissect centrosomal MTOC function. Super-resolution imaging revealed that this PCM inserts inside the lumen of the ciliary axoneme and directly nucleates the assembly of dendritic microtubules towards the cell body. Tissue-specific degradation in ciliated sensory neurons revealed a role for SPD-5 and the conserved microtubule nucleator γ-TuRC, but not SPD-2 or PCMD-1, in MTOC function at centriole-less PCM. This MTOC function was in the absence of regulation by mitotic kinases, highlighting the intrinsic ability of these proteins to drive microtubule growth and organization and further supporting a model that SPD-5 is the primary driver of MTOC function at the PCM.

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

10.1101/446088 ◽

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.

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

Journal of Cell Science ◽

10.1242/jcs.115.9.1825 ◽

2002 ◽

Vol 115 (9) ◽

pp. 1825-1835 ◽

Cited By ~ 1

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.

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Daughter Centriole Elongation Is Controlled by Proteolysis

Molecular Biology of the Cell ◽

10.1091/mbc.e09-12-1049 ◽

2010 ◽

Vol 21 (22) ◽

pp. 3942-3951 ◽

Cited By ~ 18

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.

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

Journal of Cell Science ◽

10.1242/jcs.107.2.601 ◽

1994 ◽

Vol 107 (2) ◽

pp. 601-611 ◽

Cited By ~ 1

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.

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Apical PAR complex proteins protect against epithelial assaults to create a continuous and functional intestinal lumen

10.1101/2020.10.28.359299 ◽

2020 ◽

Cited By ~ 1

Author(s):

Maria D. Sallee ◽

Melissa A. Pickett ◽

Jessica L. Feldman

Keyword(s):

Cell Division ◽

Epithelial Cells ◽

Intestinal Lumen ◽

Microtubule Organizing Center ◽

Intestinal Tube ◽

C Elegans ◽

Junctional Proteins ◽

Organizing Center ◽

Dividing Cells

ABSTRACTSustained polarity and adhesion of epithelial cells is essential for the protection of our organs and bodies, and this epithelial integrity emerges during organ development amidst numerous morphogenetic assaults. Using the developing C. elegans intestine as an in vivo model, we investigated how epithelial cells maintain integrity through cell division and elongation to build a functional tube. Live-imaging revealed that apical PAR complex proteins PAR-6/Par6 and PKC-3/aPkc remained apical during mitosis while apical microtubules and microtubule-organizing center (MTOC) proteins were transiently removed. Intestine-specific depletion of PAR-6, PKC-3, and the aPkc regulator CDC-42/Cdc42 caused persistent gaps in the apical MTOC as well as in other apical and junctional proteins after cell division and in non-dividing cells that elongated. Upon hatching, gaps coincided with luminal constrictions that blocked food, and larvae arrested and died. Thus, the apical PAR complex maintains apical and junctional continuity to construct a functional intestinal tube.

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The Centrosome and the Primary Cilium: The Yin and Yang of a Hybrid Organelle

Cells ◽

10.3390/cells8070701 ◽

2019 ◽

Vol 8 (7) ◽

pp. 701 ◽

Cited By ~ 19

Author(s):

Joukov ◽

De Nicolo

Keyword(s):

Primary Cilium ◽

Primary Cilia ◽

Microtubule Assembly ◽

Microtubule Organizing Center ◽

Environmental Signals ◽

Intracellular Signals ◽

Interphase Cells ◽

Organizing Center ◽

Pericentriolar Material ◽

Yin And Yang

Centrosomes and primary cilia are usually considered as distinct organelles, although both are assembled with the same evolutionary conserved, microtubule-based templates, the centrioles. Centrosomes serve as major microtubule- and actin cytoskeleton-organizing centers and are involved in a variety of intracellular processes, whereas primary cilia receive and transduce environmental signals to elicit cellular and organismal responses. Understanding the functional relationship between centrosomes and primary cilia is important because defects in both structures have been implicated in various diseases, including cancer. Here, we discuss evidence that the animal centrosome evolved, with the transition to complex multicellularity, as a hybrid organelle comprised of the two distinct, but intertwined, structural-functional modules: the centriole/primary cilium module and the pericentriolar material/centrosome module. The evolution of the former module may have been caused by the expanding cellular diversification and intercommunication, whereas that of the latter module may have been driven by the increasing complexity of mitosis and the requirement for maintaining cell polarity, individuation, and adhesion. Through its unique ability to serve both as a plasma membrane-associated primary cilium organizer and a juxtanuclear microtubule-organizing center, the animal centrosome has become an ideal integrator of extracellular and intracellular signals with the cytoskeleton and a switch between the non-cell autonomous and the cell-autonomous signaling modes. In light of this hypothesis, we discuss centrosome dynamics during cell proliferation, migration, and differentiation and propose a model of centrosome-driven microtubule assembly in mitotic and interphase cells. In addition, we outline the evolutionary benefits of the animal centrosome and highlight the hierarchy and modularity of the centrosome biogenesis networks.

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Microtubule nucleating sites in higher plant cells identified by an auto-antibody against pericentriolar material.

The Journal of Cell Biology ◽

10.1083/jcb.101.1.319 ◽

1985 ◽

Vol 101 (1) ◽

pp. 319-324 ◽

Cited By ~ 87

Author(s):

L Clayton ◽

C M Black ◽

C W Lloyd

Keyword(s):

Higher Plants ◽

Plant Cells ◽

Nuclear Surface ◽

Microtubule Organizing Center ◽

Higher Plant ◽

Spindle Poles ◽

Late Anaphase ◽

Organizing Center ◽

Pericentriolar Material ◽

Auto Antibody

Human scleroderma serum 5051, which is known to recognize the amorphous pericentriolar microtubule organizing center material of a variety of vertebrate cells, was found to immunostain spindle poles of meristematic higher plants from pre-prophase to late anaphase. Subsequently, during cytokinesis, staining was redistributed around the reforming telophase nuclei, but was not evident in the cytokinetic phragmoplast. At the transition between telophase and interphase, before the typical cortical interphase microtubule array was established, short microtubules radiated from the nucleus and in such cells the material recognized by 5051 was located around the daughter nuclei and not the cortex. These observations have led us to propose that the perinuclear region, or the nuclear surface, may function as a nucleation center for both spindle and interphase microtubules in higher plant cells.

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