γ-tubulin complexes and their interaction with microtubule-organizing centers

Current Opinion in Structural Biology

Volumn 9, Issue 2, 1999, Pages 250-259

  Wiese, Christiane ^a   Zheng, Yixian ^a  

^a GEOPHYSICAL LABORATORY   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

CYTOSKELETON PROTEIN; TUBULIN;

AMINO ACID SEQUENCE; CRYOELECTRON MICROSCOPY; MICROTUBULE ASSEMBLY; PRIORITY JOURNAL; PROTEIN PROTEIN INTERACTION; REVIEW; SEQUENCE HOMOLOGY;

EUKARYOTA;

EID: 0033117826     PISSN: 0959440X     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0959-440X(99)80035-9     Document Type: Article

References (75)

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32. Moritz M, Braunfeld MB, Sedat JW, Alberts B, Agard DA: Microtubule nucleation by γ-tubulin-containing rings in the centrosome. Nature 1995, 378:638-640.
33. Li Q, Joshi HC: γ-Tubulin is a minus end-specific microtubule binding protein. J Cell Biol 1995, 131:207-214.
34. Stearns T, Kirschner M: In vitro reconstitution of centrosome assembly and function: The central role of γ-tubulin. Cell 1994, 76:623-637.
35. Martin OC, Gunawardane RN, Iwamatsu A, Zheng Y: Xgrip109: A γ-tubulin-associated protein with an essential role in γ-tubulin ring complex (γTuRC) assembly and centrosome function. J Cell Biol 1998, 141:675-687. This is the first paper to describe the use of the Xenopus egg-extract-derived centrosome assembly assay [26] to test the role of γTuRC components. Xenopus γTuRC can be disrupted by exposing extracts made from crushed eggs to high salt concentrations. Functional γTuRC (as assayed by the ability of the manipulated extracts to transform sperm basal bodies into microtubule asters) is regained after salt removal. To begin to test the role of different γTuRC subunits, the authors cloned one of the subunits of the Xenopus γTuRC, Xgrip109. This protein is homologous to the yeast γ-tubulin-interacting protein, Spc98p. Disrupting the γTuRC by exposing it to a high salt buffer allowed the authors to immunoprecipitate Xgrip109 and show that γTuRC assembly is disrupted by this manipulation. In accordance with this finding, immunoprecipitating Xgrip109 also abolished centrosome formation activity.
36. Murphy SM, Urbani L, Stearns T: The mammalian γ-tubulin complex contains homologues of the yeast spindle pole body components Spc97p and Spc98p. J Cell Biol 1998, 141:663-674. This paper shows that the cytoplasmic γ-tubulin complex in mammalian tissue culture cells is similar in protein composition to the Xenopus γ-tubulin ring complex (γTuRC) (see Table 1). It further describes the cloning, by homology to Spc97p and Spc98p, of the human γ-tubulin complex proteins, hGCP2 and hGCP3. hGCP3 was also identified by Tassin et al. [37], who named it HsSpc98; the Xenopus homolog was named Xgrip109 [35]. GCP2 and GCP3 are centrosomal proteins that are also components of the human γTuRC. This paper provides evidence that the human γTuRC contains more than one copy of hGCP3. GCP2 and GCP3, along with γ-tubulin, co-sediment with microtubules in vitro.
37. Tassin AM, Celati C, Moudjou M, Bornens M: Characterization of the human homologue of the yeast Spc98p and its association with γ-tubulin. J Cell Biol 1998, 141:689-701. This paper reports the cloning by homology and the functional study of the human homolog of Spc98p (named HsSpc98 and hGCP3). Antibodies raised against HsSpc98 confirmed that it is concentrated at centrosomes and that it associates with γ-tubulin in a cytoplasmic complex, as well as at the centrosome. The HsSpc98 antibodies were also shown to inhibit microtubule nucleation from isolated centrosomes in vitro and when microinjected into cells. Furthermore, HsSpc98 and γ-tubulin antibodies stained demembranated Xenopus sperm head centrioles, which had previously been reported to be unable to nucleate microtubule asters. The authors suggest that the centrosomal complex may need to be activated somehow before it is able to nucleate microtubules.
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48. Geissler S, Pereira G, Spang A, Knop M, Soues S, Kilmartin J, Schiebel E: The spindle pole body component Spc98p interacts with the γ-tubulin-like Tub4p of Saccharomyces cerevisiae at the sites of microtubule attachment. EMBO J 1996, 15:3899-3911.
49. Knop M, Pereira G, Geissler S, Grein K, Schiebel E: The spindle pole body component Spc97p interacts with the γ-tubulin of Saccharomyces cerevisiae and functions in microtubule organization and spindle pole body duplication. EMBO J 1997, 16:1550-1564. SPC97 was first identified in a screen for dosage-dependent suppressors of a conditional lethal mutation in SPC98. SPC97 codes for an essential 97 kDa protein that localizes to both sides of the spindle pole body (SPB) by immuno-EM staining of Spc97-3HA. The authors used a number of methods to show that Spc97p exists in a complex with Spc98p and Tub4p. This complex migrates at approximately 6S on sucrose gradients. The authors provide evidence that the 6S complex contains more than one copy of Tub4p. The analysis of temperature-sensitive mutants suggest that Spc97p is involved in mitotic spindle formation, SPB duplication and SPB separation. Interestingly, microtubule nucleation and organization seemed relatively unimpaired in these mutants.
50. Knop M, Schiebel E: Spc98p and Spc97p of the yeast γ-tubulin complex mediate binding to the spindle pole body via their interaction with Spc110p. EMBO J 1997, 16:6985-6995. This very nice biochemical analysis of the Tub4p complex revealed that it is composed of Spc98, Spc97 and Tub4p in a ratio of 1:1:=2. Two-hybrid studies further showed that the Tub4p complex interacts with the N terminus of Spc110p through Spc98p and Spc97p, but not through Tub4p. Spc110p could be isolated in a complex with Spc42p, Cmd1p (the yeast calmodulin protein) and an unidentified 35 kDa protein.
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55. Nguyen T, Vinh DBN, Crawford DK, Davis TN: A genetic analysis of interactions with Spc110p reveals distinct functions of Spc97p and Spc98p, components of the yeast γ-tubulin complex. Mol Biol Cell 1998, 9:2201-2216. This paper examines the molecular details of the interaction between Spc110p and the Tub4p complex. The N terminus of Spc110p. which faces the inner plaque and is therefore the part of Spc110p that is most likely to interact with a microtubule nucleator, interacts with the Tub4p complex via Spc98p. No interaction was detected between Spc110p and Tub4p, and the interaction between Spc110p and Spc97p was shown to be dependent on Spc98p.
56. Sundberg HA, Davis TN: A mutational analysis identifies three functional regions of the spindle pole component Spc110p in Saccharomyces cerevisiae. Mol Biol Cell 1997, 8:2575-2590. The authors present genetic evidence for an interaction between Spc98p and the N terminus of Spc110p, but not the C terminus or the calmodulin-binding domain of Spc110p. This, together with [50], is the first molecular evidence that Spc110p tethers the Tub4p complex to the spindle pole body on the nuclear side.
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59. Knop M, Schiebel E: Receptors determine the cellular localization of a γ-tubulin complex and thereby the site of microtubule formation. EMBO J 1998, 17:3952-3967. Using two-hybrid technology backed up by genetic and biochemical evidence, the authors identified Spc72p as the Tub4p complex receptor on the cytoplasmic side of the yeast spindle pole body. The interaction is shown to occur through the interaction of Spc72p with Spc98p from the Tub4p complex (see also [67]).
60. Wigge PA, Jensen ON, Holmes S, Souès S, Mann M, Kilmartin JV: Analysis of the Saccharomyces spindle pole by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. J Cell Biol 1998, 141:967-977. This very important paper reports the identification of spindle pole body (SPB) components using MALDI mass spectroscopic analysis of highly enriched SPB fractions. A total of 45 (out of approximately 80) bands were analyzed, which identified 43 different proteins; 12 of these were known SPB components and 11 were novel components. Of the novel components, eight were localized to particular parts of the SPB by immuno-EM. Another eight proteins were localized to different parts of the spindle.
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63. Pereira G, Knop M, Schiebel E: Spc98p directs the yeast γ-tubulin complex into the nucleus and is subject to cell cycle-dependent phosphorylation on the nuclear side of the spindle pole body. Mol Biol Cell 1998, 9:775-793. This is the only study to date that addresses the biosynthesis of the Tub4p complex. Spc98p was discovered to contain an essential nuclear localization signal that directs the entire Tub4p complex into the nucleus. Moreover, nuclear spindle pole body associated Spc98p becomes phosphorylated in a cell-cycle-specific manner.
64. Doxsey SJ, Stein P, Evans L, Calarco PD, Kirschner M: Pericentrin, a highly conserved centrosome protein involved in microtubule organization. Cell 1994, 76:639-650.
65. Dictenberg SJ, Zimmerman W, Sparks CA, Young A, Vidair C, Zheng Y, Carrington W, Fay FS, Doxsey SJ: Pericentrin and γ-tubulin form a protein complex and are organized into a novel lattice at the centrosome. J Cell Biol 1998, 141:163-174. Using sucrose gradients, gel filtration and co-immunoprecipitation analysis, the authors show that Xenopus egg cytoplasmic γ-tubulin forms a distinct detergent-labile complex with pericentrin that is larger than the γ-tubulin ring complex (γTuRC). Combining immunofluorescence with a deconvolution algorithm, pericentrin and γ-tubulin were found to form novel, closely apposed lattice-like structures at mammalian centrosomes and at Xenopus mitotic spindle poles. Pericentrin and γ-tubulin appear to have identical assembly and disassembly kinetics during the somatic cell cycle. The size of the lattice is regulated in a cell-cycle-dependent manner. Microtubule ends were shown to be in close proximity to the lattice, suggesting a role for pericentrin in microtubule nucleation, either directly or by interacting with the γTuRC.
66. Wang PJ, Huffaker TC: Stu2p: A microtubule-binding protein that is an essential component of the yeast spindle pole body. J Cell Biol 1997, 139:1271-1280. STU2 (a suppressor of tub2) was identified as an essential, extragenic suppressor of a cold-sensitive mutation in the yeast tub2 gene, which encodes β-tubulin. Stu2p-GFP (green fluorescent protein) localizes mainly to the spindle pole body (SPB), but is also found along the microtubules. SPB localization is independent of microtubules. Biochemical evidence shows that Stu2p binds along the sides of microtubules.
67. Chen XP, Yin H, Huffaker TC: The yeast spindle pole body component Spc72p interacts with Stu2p and is required for proper microtubule assembly. J Cell Biol 1998, 141:1169-1179. Using two-hybrid analysis to identify proteins that interact with Stu2p, the authors identified Spc72p, a coiled-coil protein that lacks significant sequence homology with proteins in the database. Interaction between Spc72p and Stu2p was confirmed by co-immunoprecipitation experiments. Spc72p localizes to the outer plaque of the spindle pole body (SPB) in a microtubule-independent manner, but Spc72p does not interact with microtubules. Phenotypically, SPC72 mutant cells have defective cytoplasmic microtubules (microtubules are either absent, unusually long or not attached to the SPB), while nuclear microtubules appear to be unaffected. During mitosis, spindle elongation is inhibited. Thus, Spc72p may be involved in microtubule tethering (see also [59]).
68. dis1, which is required for sister chromatid separation, is a novel microtubule and spindle pole body-associating protein phosphorylated at the Cdc2 target sites. Genes Dev 1995, 9:1572-1585.
69. Matthews LR, Carter P, Thierry-Mieg D, Kemphues K: ZYG-9, a Caenorhabditis elegans protein required for microtubule organization and function, is a component of meiotic and mitotic spindle poles. J Cell Biol 1998, 141:1159-1168. The authors cloned the gene and characterized the protein that is responsible for the zyg-9 mutation in the worm C. elegans. ZYG-9 is localized to mitotic and meiotic spindle poles and is homologous to a family of proteins that includes Stu2p, XMAP215 and ch-TOG. RNA interference experiments show that ZYG-9 is required for the formation of long microtubules.
70. Charasse S, Schroeder M, Gauthier-Rouviere C, Ango F, Cassimeris L, GarD D, Larroque C: The TOGp is a new human microtubule-associated protein homologous to the Xenopus XMAP215. J Cell Sci 1998, 111:1371-1383. The recently discovered colonie, hepatic tumor overexpressed gene (ch-TOG) was shown to encode a 218 kDa human homolog of the Xenopus XMAP215 protein. The ch-TOG antigen decorated centrosomes and spindles in a cell-cycle-specific manner, co-sedimented with taxol-stabilized microtubules (MTs) in vitro and promoted MT assembly.
71. Gard DL, Kirschner MW: A microtubule-associated protein from Xenopus eggs that specifically promotes assembly at the plus-end. J Cell Biol 1987, 105:2203-2215.
72. Vasquez RJ, Gard D, Cassimeris L: XMAP from Xenopus eggs promotes rapid plus end assembly of microtubules and rapid microtubule polymer turnover. J Cell Biol 1994, 127:985-993.
73. Schnackenberg BJ, Khodjakov A, Rieder CL, Palazzo RE: The disassembly and reassembly of functional centrosomes in vitro. Proc Natl Acad Sci USA 1998, 95:9295-9300. This paper describes the effects of the reversible salt treatment of centrosomes (see also [27]). Extracting Spisula centrosomes with 1 M potassium iodide (Kl) abolishes their microtubule nucleating activity and results in a centrosome 'ghost' (termed 'centromatrix' by the authors) - a complex network of 12-15 nm fibers. Salt treatment also removes centrioles from the 'centromatrix', as well as destroying γ-tubulin-containing ring structures (approximately 25 nm in diameter) and extracting γ-tubulin. Microtubule nucleating activity, γ-tubulin and the 25 nm ring structures can be recovered by incubating the 'centromatrix' with oocyte extracts. Recovery is independent of microtubules, nucleotides or divalent cations.
74. Tassin AM, Celati C, Paintrand M, Bornens M: Identification of an Spc110p-related protein in vertebrates. J Cell Sci 1997, 110:2533-2545. The authors used antibodies directed against the yeast Spc110p, which interacts with the Tup4p complex on the nuclear side of the spindle pole body, to probe mammalian centrosomes. Antibodies against Spc110p decorated mammalian centrosomes, inhibited microtubule nucleation from isolated human centrosomes and cross-reacted with an approximately 121 kDa band that co-immunoprecipitated with an approximately 25S γ-tubulin complex partially purified from Xenopus egg extracts. These results suggest that vertebrate centrosomes contain at least one protein that is immunologically related to Spc110p.
75. Geissler S, Siegers K, Schiebel E: A novel protein complex promoting formation of functional α- and γ-tubulin. EMBO J 1998, 17:952-966. Using synthetic lethality with a mutated γ-tubulin gene in S. cerevisiae as a criterion to screen for γ-tubulin-interacting proteins, the authors identified a set of proteins that are related to one another and appear to be involved in the proper formation and folding of both γ- and α-tubulin. These proteins are named GIMs, for genes involved in microtubule biogenesis. GIMs form cytoplasmic multiprotein complexes that are not associated with the spindle pole body.