Gamma-Tubulin is a phylogenetically conserved component of microtubule-organizing centers that is essential for viability and microtubule function. Expression of a human gamma-tubulin cDNA restores viability and a near-normal growth rate to cells of S. pombe lacking endogenous gamma-tubulin. Immunofluorescence microscopy shows that these cells contain normal mitotic spindles and interphase microtubule arrays, and that human gamma-tubulin, like S. pombe gamma-tubulin, localizes to spindle pole bodies, the fungal microtubule-organizing centers. These results demonstrate that human gamma-tubulin functions in fission yeast, and suggest that in spite of the great morphological differences between the microtubule-organizing centers of humans and fission yeasts, gamma-tubulin is likely to perform the same tasks in both. They suggest, moreover, that the proteins that interact with gamma-tubulin must also be conserved, including, most obviously, microtubule-organizing center proteins. A fivefold overexpression of S. pombe gamma-tubulin causes no reduction in growth rates or alteration of microtubule organization. It is throught that excess gamma-tubulin is maintained in the cytoplasm in a form incapable of nucleating microtubule assembly (Horio, 1994)

Pericentrin and gamma-tubulin are integral centrosome proteins that play a role in microtubule nucleation and organization. The relationship between these proteins was studied in the cytoplasm and at the centrosome. In extracts prepared from Xenopus eggs, the proteins are part of a large complex as demonstrated by sucrose gradient sedimentation, gel filtration and coimmunoprecipitation analysis. The pericentrin-gamma-tubulin complex is distinct from the previously described gamma-tubulin ring complex (gamma-TuRC), because purified gamma-TuRC fractions do not contain detectable pericentrin. When assembled at the centrosome, the two proteins remain in close proximity, as shown by fluorescence resonance energy transfer. The three-dimensional organization of the centrosome-associated fraction of these proteins was determined using an improved immunofluorescence method. This analysis reveals a novel reticular lattice that is conserved from mammals to amphibians, and is organized independent of centrioles. The lattice changes dramatically during the cell cycle, enlarging from G1 until mitosis, then rapidly disassembling as cells exited mitosis. In cells colabeled to detect centrosomes and nucleated microtubules, lattice elements appear to contact the minus ends of nucleated microtubules. These results indicate that pericentrin and gamma-tubulin assemble into a unique centrosome lattice that represents the higher-order organization of microtubule nucleating sites at the centrosome (Dictenberg, 1998).

The localization of RNAs at the vegetal cortex in Xenopus oocytes is a complex process, involving at least two different pathways. The early, or messenger transport organizer (METRO), pathway localizes RNAs such as Xlsirts, Xcat2 and Xwnt11 during stages 1 and 2 of oogenesis, while the late pathway localizes RNAs such as Vg1 during stages 2-4. The onset of Vg1 localization is characterized by its microtubule-independent binding to a subdomain of the endoplasmic reticulum (ER). The formation of this unique ER structure is intimately associated with the movement of the mitochondrial cloud toward the vegetal cortex. In addition, the mitochondrial cloud contains a gamma-tubulin-positive structure that may function as a microtubule organizing center for establishing microtubule tracks for Vg1 localization. These data support, although they do not prove, a model in which the development of the late pathway machinery relies on the prior functioning of the early pathway (Kloc, 1998).

The in vivo function of gamma-tubulin in mammalian cells was investigated using a synthetic peptide to generate a polyclonal antibody that binds to a highly conserved segment of gamma-tubulin. After microinjection into cultured mammalian cells, immunofluorescence localization reveals that this antibody binds to native centrosomes at all phases of the cell cycle. In the presence of the gamma-tubulin antibody, microtubules fail to regrow into cytoplasmic arrays after depolymerization induced by nocodazole or cold. Furthermore, cells injected immediately before or during mitosis fail to assemble a functional spindle. Thus in vivo gamma-tubulin is required for microtubule nucleation throughout the mammalian cell cycle (Joshi, 1992).

Animal cells undergoing cytokinesis form the midbody, an inter-cellular bridge containing two bundles of microtubules interdigitated at their plus ends. Polyclonal antibodies raised against three specific amino acid sequences of gamma-tubulin specifically stained the centrosome in interphase, the spindle poles in all stages of mitosis, and the extremities of the midbody in mammalian cells. No gamma-tubulin could be detected in the interzone during anaphase and early telophase. Material containing gamma-tubulin first appears in the two daughter cells on each side of the division plane in late telophase, and for one hour after metaphase, accumulates transiently at the minus ends of the two microtubule bundles constituting the midbody. Micro-injection of gamma-tubulin antibodies into anaphase cells prevents the subsequent formation of the microtubule bundles between the two daughter cells. In contrast to previous views, these observations suggest that the microtubules constituting the midbody may be nucleated on special microtubule organizing centers, active during late telophase only, and assembled on each side of the dividing plane between the daughter cells (Julian, 1993).

To test whether gamma-tubulin mediates the nucleation of microtubule assembly in vivo, and co-assembles with alpha- and beta-tubulins into microtubules or self-assembles into macro-molecular structures, the expression of gamma-tubulin was elevated in the cell cytoplasm. In most cells, overexpression of gamma-tubulin causes a dramatic reorganization of the cellular microtubule network. When overexpressed, gamma-tubulin causes ectopic nucleation of microtubules that are not associated with the centrosome. In a fraction of cells, gamma-tubulin self-assembles into novel tubular structures with a diameter of approximately 50 nm (named gamma-tubules). Furthermore, unlike microtubules, gamma-tubules are resistant to cold or drug induced depolymerization. These data provide evidence that gamma-tubulin can cause nucleation of microtubule assembly and can self-assemble into novel tubular structures (Shu, 1995).

The highly conserved protein gamma-tubulin is required for microtubule nucleation in vivo. When viewed in the electron microscope, a highly purified gamma-tubulin complex from Xenopus consisting of at least seven different proteins is seen to have an open ring structure. This complex acts as an active microtubule-nucleating unit that can cap the minus ends of microtubules in vitro (Zheng, 1995).

Previous studies indicate that gamma tubulin ring complex (gammaTuRC) can nucleate microtubule assembly and may be important in centrosome formation. gammaTuRC contains approximately eight subunits, termed Xenopus gamma ring proteins (Xgrips), in addition to gamma tubulin. One gammaTuRC subunit, Xgrip109, is a highly conserved protein, with homologs present in yeast, rice, flies, zebrafish, mice, and humans. The yeast Xgrip109 homolog, Spc98, is a spindle-pole body component that interacts with gamma tubulin. In vertebrates, Xgrip109 identifies two families of related proteins. Xgrip109 and Spc98 have more homology to one family than the other. Xgrip109 is a centrosomal protein that directly interacts with gamma tubulin. A complementation assay for centrosome formation has been developed using demembranated Xenopus sperm and Xenopus egg extract. Using this assay, it has been shown that Xgrip109 is necessary for the reassembly of salt-disrupted gammaTuRC and for the recruitment of gamma tubulin to the centrosome. Xgrip109, therefore, is essential for the formation of a functional centrosome (Martin, 1998).

gamma-Tubulin is a universal component of microtubule organizing centers, where it is believed to play an important role in the nucleation of microtubule polymerization. gamma-Tubulin also exists as part of a cytoplasmic complex whose size and complexity varies in different organisms. To investigate the composition of the cytoplasmic gamma-tubulin complex in mammalian cells, cell lines stably expressing epitope-tagged versions of human gamma-tubulin were made. The epitope-tagged gamma-tubulins expressed in these cells localize to the centrosome and are incorporated into the cytoplasmic gamma-tubulin complex. Immunoprecipitation of this complex identifies at least seven proteins, with calculated molecular weights of 48, 71, 76, 100, 101, 128, and 211 kD. The 100- and 101-kD components of the gamma-tubulin complex have been identified as homologs of the yeast spindle pole body proteins Spc97p and Spc98p; the corresponding human proteins have been named hGCP2 and hGCP3. Sequence analysis reveals that these proteins are not only related to their respective homologs, but are also related to each other. GCP2 and GCP3 colocalize with gamma-tubulin at the centrosome, cosediment with gamma-tubulin in sucrose gradients, and coimmunoprecipitate with gamma-tubulin, indicating that they are part of the gamma-tubulin complex. The conservation of a complex involving gamma-tubulin, GCP2, and GCP3 from yeast to mammals suggests that structurally diverse microtubule organizing centers such as the yeast spindle pole body and the animal centrosome share a common molecular mechanism for microtubule nucleation (Murphy, 1998).

A novel human protein with a molecular mass of 55 kD, designated RanBPM, was isolated with the two-hybrid method, using Ran as a bait. Mouse and hamster RanBPM possessed a polypeptide identical to the human one. Furthermore, Saccharomyces cerevisiae was found to have a gene, YGL227w, the COOH-terminal half of which is 30% identical to RanBPM. Anti-RanBPM antibodies reveal that RanBPM is localized within the centrosome throughout the cell cycle. Overexpression of RanBPM produces multiple spots that are colocalized with gamma-tubulin and act as ectopic microtubule nucleation sites, resulting in a reorganization of the microtubule network. RanBPM cosediments with the centrosomal fractions by sucrose-density gradient centrifugation. The formation of microtubule asters is inhibited not only by anti-RanBPM antibodies, but also by nonhydrolyzable GTP-Ran. Indeed, RanBPM specifically interacts with GTP-Ran in a two-hybrid assay. The central part of asters stained by anti-RanBPM antibodies or by the mAb to gamma-tubulin is faded by the addition of GTPgammaS-Ran, but not by the addition of anti-RanBPM antibodies. These results provide evidence that the Ran-binding protein, RanBPM, is involved in microtubule nucleation, thereby suggesting that Ran regulates the centrosome through RanBPM (Nakamura, 1998).

Epithelial polarity, the ability of simple epithelial cells to become asymmetric, with distinct apical and basolateral domains in the plasma membrane, is a puzzling, as yet unsolved case of how cells can generate different subdomains. There is a consensus that epithelial cells sort out newly synthesized membrane proteins at the trans-Golgi network and recycle membrane proteins at an intermediate endosomal compartment. The vesicular traffic originating from these sorting compartments is probably directed by cytoskeletal components to their final destinations in the apical or basolateral domains and retained in place. Although epithelial cells can deliver polarized proteins without organized microtubules, several lines of evidence support the idea that microtubule based motors participate in the movement of exocytic vesicles, at least along part of their pathway. Moreover, in well-differentiated simple epithelia, microtubules are organized in the apico-basal axis with their minus ends toward the apical side. This organization is thought to participate in the polarization of organelles in the cytoplasm and to contribute to the polarized vesicular traffic. This peculiar arrangement of the microtubules must result from a polarized distribution of microtubule organizing centers (MTOC) under the apical domain. In fact, centrosomal structures distribute under the apical membrane in a number of simple epithelium cells in interphase, both ciliated and nonciliated. Interestingly, when epithelial cells enter mitosis, the centrosomes move toward the lateral domains, organizing the mitotic spindle always perpendicular to the apico-basal axis, and then move back to the apical domain when the cells complete mitosis. The significance of this elaborate redistribution of centrosomes is currently unknown. It is especially intriguing since apical (interphasic) centrosomes retain their capability to act as MTOC while most of the microtubules are organized by noncentrosomal MTOCs, diffusely distributed under the apical domain (Salas, 1999 and references).

A network of intermediate filaments (IF) underlying the apical membrane of epithelial cells contains cytokeratin (CK) 19 IF. These filaments are attached to a subpopulation of apical membrane proteins. Furthermore, this IF network, equivalent to the terminal web, is observed in a variety of epithelial cells with or without brush-border. Using antisense oligonucleotides, CK19 could be transiently downregulated in a fraction of CACO-2 epithelial cells. Those cells display a characteristic phenotype, with abolished apical F-actin (while the rest of cortical F-actin is normal), distinctive changes in the polarization of membrane proteins, and disorganization of the apical, but not basal, network of microtubules. The latter observation, together with the reports of an apical distribution of MTOC, prompted a hypothesis that the apical IF may be responsible for the polarized distribution of gamma-tubulin containing structures in simple epithelia. It is generally accepted that gamma-tubulin is an essential component of MTOC. It remains associated with the centrosomes even when the microtubules are depolymerized. Approximately 50% of the gamma-tubulin is in a soluble form, part of a 28S complex, while the rest is insoluble, presumably associated with centrosomes. Therefore, the possibility of MTOC binding to apical IF was analyzed, by studying the attachment of insoluble gamma-tubulin to CKs (Salas, 1999 and references).

The apical distribution of gamma-tubulin containing structures (potential microtubule-organizing centers) in CACO-2 cells was confirmed and perfect colocalization of centrosomes was demonstrated and nearly 50% of noncentrosomal gamma-tubulin with apical intermediate filaments, but not with apical F-actin. Furthermore, the antisense-oligonucleotide mediated downregulation of cytokeratin 19, using two different antisense sequences, is more efficient than anticytoskeletal agents to delocalize centrosomes. Electron microscopy colocalization suggests that binding occurs at the outer boundary of the pericentriolar material. Type I cytokeratins 18 and 19 present in these cells specifically coimmunoprecipitates in multi-protein fragments of the cytoskeleton with gamma-tubulin. The size and shape of the fragments, visualized at the EM level, indicate that physical trapping is an unlikely explanation for this result. Drastic changes in the extraction protocol do not affect coimmunoprecipitation. These results from three independent techniques indicate that insoluble gamma-tubulin containing structures are attached to apical intermediate filaments (Salas, 1999).

The centrosome organizes microtubules (which are made up of alpha-tubulin and beta-tubulin) and contains centrosome-bound gamma-tubulin, which is involved in microtubule nucleation. Two new human tubulins have been identified and they are shown to be are associated with the centrosome. One is a homolog of the Chlamydomonas delta-tubulin Uni3, and the other is a new tubulin, which has been named epsilon-tubulin. Localization of delta-tubulin and epsilon-tubulin to the centrosome is independent of microtubules, and the patterns of localization are distinct from each other and from that of gamma-tubulin. delta-Tubulin is found in association with the centrioles, whereas epsilon-tubulin localizes to the pericentriolar material. epsilon-Tubulin exhibits a cell-cycle-specific pattern of localization, first associating with only the older of the centrosomes in a newly duplicated pair and later associating with both centrosomes. epsilon-Tubulin thus distinguishes the old centrosome from the new at the level of the pericentriolar material, indicating that there may be a centrosomal maturation event that is marked by the recruitment of epsilon-tubulin (Chang, 2000).

The first delta-tubulin described, Uni3 from Chlamydomonas, was identified from a mutant with defects in flagellar biogenesis and in the structure of the triplet microtubules of the basal body/centriole. The localization of human delta-tubulin to the region around the centrioles points to a potentially similar role for this protein in animal cells; however, the prominent intercentriolar localization of delta-tubulin suggests other possible functions as well. For example, in Clamydomonas, fibers linking the two centrioles within the centrosome have been found, and in vitro studies of centrosome duplication indicate that separation of the centrioles, and thus dissolution of the fibers, may be an early regulated step in the process. The unusual localization pattern of delta-tubulin resembles in several respects that observed for Skp1, a component of the SCF ubiquitin ligase, which is required for centriole separation (Chang, 2000).

epsilon-Tubulin has not been described previously: the only clue to its function comes from the cell-cycle-specific localization. Given that the single centrosome of G1 cells begins the cycle having epsilon-tubulin, the simplest interpretation of the differential localization in S and early G2 is that the pericentriolar material of one centrosome is different from that of the other after duplication. The newer centrosome, defined by the age of the parental centriole within the centrosome, acquires epsilon-tubulin only after some maturation process. This maturation is independent of the acquisition of nucleating capacity by the new centrosome, but might reflect some other function that varies with the cell cycle. The differential localization of epsilon-tubulin to old and new centrosomes is particularly interesting in light of the finding (Lange, 1995) that cenexin is a protein that associates only with the older centriole within a centrosome. Centriole distribution is semi-conservative; after centriole separation, the newer centriole of the original pair does not acquire cenexin until the G2/M transition, roughly at the same time that the new centrosome has acquired a full complement of epsilon-tubulin. The epsilon-tubulin results also indicate that duplication of the centrosome does not involve merely partitioning of the existing pericentriolar material around the two pairs of centrioles resulting from duplication, but rather that the centrosome containing the newer centriole also has 'new' pericentriolar material. The functional consequences of the difference between the duplicated centrosomes remain to be determined, but an attractive possibility is that it might be involved in the regulation of a cell-cycle-specific function, such as centrosome duplication, or cell-cycle-specific recruitment of other molecules to the centrosome (Chang, 2000).

This study shows that cytoplasmic dynein mediates assembly of pericentrin and gamma tubulin onto centrosomes. Centrosome assembly is important for mitotic spindle formation and if defective may contribute to genomic instability in cancer. In somatic cells, centrosome assembly of two proteins involved in microtubule nucleation, pericentrin and gamma tubulin, is inhibited in the absence of microtubules. A more potent inhibitory effect on centrosome assembly of these proteins is observed after specific disruption of the microtubule motor cytoplasmic dynein by microinjection of dynein antibodies or by overexpression of the dynamitin subunit of the dynein binding complex dynactin. Consistent with these observations is the ability of pericentrin to cosediment with taxol-stabilized microtubules in a dynein- and dynactin-dependent manner. Centrosomes in cells with reduced levels of pericentrin and gamma tubulin have a diminished capacity to nucleate microtubules. In living cells expressing a green fluorescent protein-pericentrin fusion protein, green fluorescent protein particles containing endogenous pericentrin and gamma tubulin move along microtubules at speeds of dynein and dock at centrosomes. In Xenopus extracts where gamma tubulin assembly onto centrioles can occur without microtubules, assembly is enhanced in the presence of microtubules and inhibited by dynein antibodies. From these studies it is concluded that pericentrin and gamma tubulin are novel dynein cargoes that can be transported to centrosomes on microtubules and whose assembly contributes to microtubule nucleation (Young, 2000).

Based on this data, a model is proposed for the assembly of microtubule nucleating proteins. In this model, pericentrin binds to dynein through the light intermediate chain and to the gamma tubulin ring complex (gamma TuRC) through specific subunits of this complex. Dynein would mediate binding of the large pericentrin-gamma TuRC complex to microtubules and direct transport of the complex to centrosomes. At the centrosome, pericentrin-gamma TuRC complexes would be anchored, whereas dynein could be released for additional rounds of transport or anchored to perform additional roles. Dynactin may facilitate microtubule association or processivity of dynein and may contribute to centrosomal anchoring of gamma tubulin. This work raises the possibility that pericentrin mediates centrosome and spindle function through dynein-dependent assembly of microtubule nucleating complexes and other activities (Young, 2000).

There is now good evidence for microtubule-dependent and microtubule-independent mechanisms for recruitment of proteins onto centrosomes. These studies support the idea that dynein-mediated and passive diffusion mechanisms represent parallel pathways for centrosome assembly. It is possible that one pathway predominates over the other in certain biological systems or at different stages of the cell cycle. In embryonic systems, for example, high levels of centrosome proteins may be sufficient to drive the initial stages of microtubule-independent recruitment onto centrioles, although dynein-mediated transport becomes a major contributor at later times. Alternative mechanisms could also account for centrosome protein recruitment. Spontaneously assembled microtubules could be capped by gamma tubulin (and pericentrin) complexes, and these small microtubule fragments could be transported toward the minus ends of microtubules by dynein as described during spindle assembly in Xenopus extracts. These data do not distinguish between this microtubule fragment mechanism and the model presented in this paper in which presumably inactive centrosome proteins are transported to centrosomes and become active for microtubule nucleating activity. Another possibility is that centrosome-nucleated microtubules are released but remain tethered to the centrosome, perhaps through an interaction with dynactin, and they provide new minus ends for binding of gamma tubulin-pericentrin complexes after passive diffusion to these sites. Although this mechanism could account for the microtubule dependency of centrosome protein recruitment, it is inconsistent with kinetic data showing directed movement of GFP-pericentrin toward centrosomes (Young, 2000).

The topography of microtubule assembly events during meiotic maturation of animal oocytes demands tight spatial control and temporal precision. To better understand what regulates the timing and location of microtubule assembly, synchronously maturing mouse oocytes were evaluated with respect to gamma-tubulin, pericentrin, and total tubulin polymer fractions at specific stages of meiotic progression. gamma-Tubulin remains associated with cytoplasmic centrosomes through diakinesis of meiosis-1. Following chromatin condensation and perinuclear centrosome aggregation, gamma-tubulin relocates to a nuclear lamina-bounded compartment in which meiosis-1 spindle assembly occurs. gamma-Tubulin is stably associated with the meiotic spindle from prometaphase-1 through to anaphase-2, but also exhibits cell cycle-specific relocalization to cytoplasmic centrosomes. Specifically, anaphase onset of both meiosis-1 and -2 is characterized by the concomitant appearance of gamma-tubulin and microtubule nucleation in subcortical centrosomes. Brief pulses of taxol applied at specific cell cycle stages enhances detection of gamma-tubulin compartmentalization, consistent with a gamma-tubulin localization-dependent spatial restriction of microtubule assembly during meiotic progression. In addition, a taxol pulse during meiotic resumption impairs subsequent gamma-tubulin sorting, resulting in monopolar spindle formation and cell cycle arrest in meiosis-1; despite cell cycle arrest, polar body extrusion occurred roughly on schedule. Therefore, sorting of gamma-tubulin is involved in both the timing of location of meiotic spindle assembly as well as the coordination of karyokinesis and cytokinesis in mouse oocytes (Combelles, 2001).

Since gamma-tubulin is a known rate-limiting determinant of microtubule nucleation, it was postulated that redistribution of gamma-tubulin could underlie maturation-associated changes in microtubule patterning. The main findings of this study include: (1) redistribution of gamma-tubulin from predominantly cytoplasmic centrosomes to a nuclear compartment delimited by a persistent nuclear lamina; (2) sorting of gamma-tubulin between cytoplasmic and nuclear (spindle) compartments during meiotic progression based on the reappearance of gamma-tubulin in cytoplasmic centrosomes during anaphase of meiosis-1 and -2, and (3) functional uncoupling of karyokinesis and cytokinesis as a result of impaired sorting of gamma-tubulin after application of brief pulses of taxol. Together, these observations suggest that modifications in microtubule patterning during meiotic maturation are due to temporally and spatially regulated changes in gamma-tubulin distribution (Combelles, 2001).

Microtubule assembly is initiated by the gamma-tubulin ring complex (gamma-TuRC). In yeast, the microtubule is nucleated from gamma-TuRC anchored to the amino-terminus of the spindle pole body component Spc110p, which interacts with calmodulin (Cmd1p) at the carboxy-terminus. However, mammalian protein that anchors gamma-TuRC remains to be elucidated. A giant coiled-coil protein, CG-NAP (centrosome and Golgi localized PKN-associated protein), has been localized to the centrosome via the carboxyl-terminal region. This region interacts with calmodulin by yeast two-hybrid screening, and it shares high homology with the carboxyl-terminal region of kendrin, another centrosomal coiled-coil protein. The amino-terminal region of either CG-NAP or kendrin indirectly associates with gamma-tubulin through binding with gamma-tubulin complex protein 2 (GCP2) and/or GCP3. Furthermore, endogenous CG-NAP and kendrin coimmunoprecipitate with each other and with endogenous GCP2 and gamma-tubulin, suggesting that CG-NAP and kendrin form complexes and interact with gamma-TuRC in vivo. These proteins localize to the center of microtubule asters nucleated from isolated centrosomes. Pretreatment of the centrosomes by antibody to CG-NAP or kendrin moderately inhibits the microtubule nucleation; moreover, the combination of these antibodies results in stronger inhibition. These results imply that CG-NAP and kendrin provide sites for microtubule nucleation in the mammalian centrosome by anchoring gamma-TuRC (Takahashi, 2002).

Microtubule nucleation is the best known function of centrosomes. Centrosomal microtubule nucleation is mediated primarily by gamma tubulin ring complexes (gamma TuRCs). However, little is known about the molecules that anchor these complexes to centrosomes. This study, carried out with Xenopus extracts, shows that the centrosomal coiled-coil protein pericentrin anchors gammaTuRCs at spindle poles through an interaction with gamma tubulin complex proteins 2 and 3 (GCP2/3). Pericentrin silencing by small interfering RNAs in somatic cells disrupts gamma tubulin localization and spindle organization in mitosis but has no effect on gamma tubulin localization or microtubule organization in interphase cells. Similarly, overexpression of the GCP2/3 binding domain of pericentrin disrupts the endogenous pericentrin-gammaTuRC interaction and perturbs astral microtubules and spindle bipolarity. When added to Xenopus mitotic extracts, this domain uncouples gammaTuRCs from centrosomes, inhibits microtubule aster assembly, and induces rapid disassembly of preassembled asters. All phenotypes are significantly reduced in a pericentrin mutant with diminished GCP2/3 binding and are specific for mitotic centrosomal asters, since little effect was observed on interphase asters or on asters assembled by the Ran-mediated centrosome-independent pathway. Additionally, pericentrin silencing or overexpression induces G2/antephase arrest followed by apoptosis in many but not all cell types. It is concluded that pericentrin anchoring of gamma tubulin complexes at centrosomes in mitotic cells is required for proper spindle organization and that loss of this anchoring mechanism elicits a checkpoint response that prevents mitotic entry and triggers apoptotic cell death (Zimmerman, 2004).

The spindle assembly checkpoint guards the fidelity of chromosome segregation. It requires the close cooperation of cell cycle regulatory proteins and cytoskeletal elements to sense spindle integrity. The role of the centrosome, the organizing center of the microtubule cytoskeleton, in the spindle checkpoint is unclear. This study found that the molecular requirements for a functional spindle checkpoint included components of the large gamma-tubulin ring complex. However, their localization at the centrosome and centrosome integrity were not essential for this function. Thus, the spindle checkpoint can be activated at the level of microtubule nucleation (Muller, 2006).