Characterization of Two Related <i>Drosophila</i> γ-tubulin Complexes that Differ in Their Ability to Nucleate Microtubules

Oegema, Karen; Wiese, Christiane; Martin, Ona C.; Milligan, Ronald A.; Iwamatsu, Akihiro; Mitchison, Timothy J.; Zheng, Yixian

doi:10.1083/jcb.144.4.721

Cited by 292 publications

(426 citation statements)

References 45 publications

Supporting

Mentioning

395

Contrasting

Unclassified

Order By: Relevance

“…All fluorescent double in situs were, therefore, carried out using reciprocal probe combinations (i.e., hunchback-DIG probe þ pangolin-BIO probe and hunchback-BIO probe þ pangolin-DIG probe). Gene fragments were amplified using the following primers: T. Protein immunostainings were carried out according to established protocols (Patel, 1994 Oegema et al, 1999). Factin staining was carried out using Phaloidin-FITC (Sigma, Cat.…”

Section: Experimental Procedures In Situ Hybridization and Antibody Smentioning

confidence: 99%

Early asymmetries in maternal transcript distribution associated with a cortical microtubule network and a polar body in the beetle Tribolium castaneum

Peel

Averof

2010

Developmental Dynamics

View full text Add to dashboard Cite

The localization of maternal mRNAs during oogenesis plays a central role in axial specification in some insects. Here we describe a polar body-associated asymmetry in maternal transcript distribution in preblastoderm eggs of the beetle Tribolium castaneum. Since the position of the polar body marks the future dorsal side of the embryo, we have investigated whether this asymmetry in mRNA distribution plays a role in dorsal-ventral axis specification. Whilst our results suggest polar body-associated transcripts do not play a significant role in specifying the DV axis, at least during early embryogenesis, we do find that the polar body is closely associated with a cortical microtubule network (CMN), which may play a role in the localization of transcripts during oogenesis. Transcripts of the gene T.c.pangolin co-localize with the CMN at the time of their anterior localization during oogenesis and their anterior localization is disrupted by the microtubule-depolymerizing agent colcemid. Developmental Dynamics 239:2875-2887,

show abstract

Section: Experimental Procedures In Situ Hybridization and Antibody Smentioning

confidence: 99%

Early asymmetries in maternal transcript distribution associated with a cortical microtubule network and a polar body in the beetle Tribolium castaneum

Peel

Averof

2010

Developmental Dynamics

View full text Add to dashboard Cite

show abstract

“…(43,44) In α tubulin, which catalyzes the GTP of β tubulin below, there is a conservative change to E254, and this glutamate is also essential for catalysis. In β tubulin, which does not catalyze hydrolysis, this residue is K and, in γ tubulin, which has low or negligible GTP hydrolysis, (35,45) it is G or S.…”

Section: Amino Acids Highly Conserved From Ftsz To Tubulin Are Those mentioning

confidence: 99%

Evolution of the cytoskeleton

2007

View full text Add to dashboard Cite

SummaryThe eukaryotic cytoskeleton appears to have evolved from ancestral precursors related to prokaryotic FtsZ and MreB. FtsZ and MreB show 40−50% sequence identity across different bacterial and archaeal species. Here I suggest that this represents the limit of divergence that is consistent with maintaining their functions for cytokinesis and cell shape. Previous analyses have noted that tubulin and actin are highly conserved across eukaryotic species, but so divergent from their prokaryotic relatives as to be hardly recognizable from sequence comparisons. One suggestion for this extreme divergence of tubulin and actin is that it occurred as they evolved very different functions from FtsZ and MreB. I will present new arguments favoring this suggestion, and speculate on pathways. Moreover, the extreme conservation of tubulin and actin across eukaryotic species is not due to an intrinsic lack of variability, but is attributed to their acquisition of elaborate mechanisms for assembly dynamics and their interactions with multiple motor and binding proteins. A new structure-based sequence alignment identifies amino acids that are conserved from FtsZ to tubulins. The highly conserved amino acids are not those forming the subunit core or protofilament interface, but those involved in binding and hydrolysis of GTP. Historical introduction Discoveries of the prokaryotic cytoskeletonBefore 1990, the cytoskeleton was thought to have evolved only in eukaryotes. Relatives or homologs of actin, tubulin or intermediate filaments were unknown in bacteria or archaea. There were sporadic reports of candidates for bacterial actin and tubulin in the 1970s and 1980s but, apart from some intriguing images of microtubules in certain bacteria,(1) these turned out to be wrong and/or were not followed up.The first suggestions of a bacterial homolog of tubulin appeared in 1992. Three groups independently discovered that the bacterial cell division protein FtsZ bound and hydrolyzed GTP, as does tubulin, and had a seven-amino-acid sequence, GGGTGTG, virtually identical to the "tubulin signature sequence".(2-4) Mukherjee and Lutkenhaus(37) then extended the sequence alignment to find a dozen additional amino acids that were completely conserved in FtsZ and in α, β and γ tubulins. They also showed that FtsZ assembled in vitro into filamentous structures. Erickson et al. (5) found that FtsZ assembled into protofilaments that could adopt two conformations, straight and curved, similar to the straight protofilaments of the microtubule wall and the tubulin rings that peel away from microtubules during disassembly. Any question of homology was resolved when tubulin and FtsZ were found to have virtually identical structures at the level of protein folding.(6,7)If one looks for bacterial relatives of tubulin or actin using a simple computer search for sequence similarity, nothing will be found. In 1992 Bork et al.(8) used a sophisticated structurebased sequence alignment, and they discovered that bacteria did have genes related to actin. Actin is...

show abstract

“…␥ TuRCs in vertebrate cells and Drosophila contain orthologues of the three yeast proteins (␥ tubulin and ␥ complex proteins 2 and 3 [GCP2, 3]) as well as several additional components (Zheng et al, 1995;Martin et al, 1998;Moritz et al, 1998;Murphy et al, 1998Murphy et al, , 2001Oegema et al, 1999; reviewed in Job et al, 2003). In vertebrates, the centrosome protein pericentrin (pericentrin A) forms a large complex with ␥ tubulin in the cytoplasm, and the two proteins are also in proximity at the centrosome (Dictenberg et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

“…Spc97p and Spc98p mediate binding of the complex to Spc110p and Spc72p (Knop and Schiebel, 1997;Knop and Schiebel, 1998;Nguyen et al, 1998). Although there is no apparent homology between their SPC97/98 interacting domains, chimeras formed by fusing the binding domain of one with the localization domain of the other can rescue knockouts of the proteins encoding the localization domains, suggesting that the two binding domains are functionally homologous (Knop and Schiebel, 1998).␥ TuRCs in vertebrate cells and Drosophila contain orthologues of the three yeast proteins (␥ tubulin and ␥ complex proteins 2 and 3 [GCP2, 3]) as well as several additional components (Zheng et al, 1995;Martin et al, 1998;Moritz et al, 1998;Murphy et al, 1998Murphy et al, , 2001Oegema et al, 1999; reviewed in Job et al, 2003). In vertebrates, the centrosome protein pericentrin (pericentrin A) forms a large complex with ␥ tubulin in the cytoplasm, and the two proteins are also in proximity at the centrosome (Dictenberg et al, 1998).…”

mentioning

confidence: 99%

Mitosis-specific Anchoring of γ Tubulin Complexes by Pericentrin Controls Spindle Organization and Mitotic Entry

Zimmerman

Sillibourne

Rosa

et al. 2004

MBoC

274

258

View full text Add to dashboard Cite

Microtubule nucleation is the best known function of centrosomes. Centrosomal microtubule nucleation is mediated primarily by ␥ tubulin ring complexes (␥ TuRCs). However, little is known about the molecules that anchor these complexes to centrosomes. In this study, we show that the centrosomal coiled-coil protein pericentrin anchors ␥ TuRCs at spindle poles through an interaction with ␥ tubulin complex proteins 2 and 3 (GCP2/3). Pericentrin silencing by small interfering RNAs in somatic cells disrupted ␥ tubulin localization and spindle organization in mitosis but had no effect on ␥ tubulin localization or microtubule organization in interphase cells. Similarly, overexpression of the GCP2/3 binding domain of pericentrin disrupted the endogenous pericentrin-␥ TuRC interaction and perturbed astral microtubules and spindle bipolarity. When added to Xenopus mitotic extracts, this domain uncoupled ␥ TuRCs from centrosomes, inhibited microtubule aster assembly, and induced rapid disassembly of preassembled asters. All phenotypes were significantly reduced in a pericentrin mutant with diminished GCP2/3 binding and were specific for mitotic centrosomal asters as we observed little effect on interphase asters or on asters assembled by the Ran-mediated centrosome-independent pathway. Additionally, pericentrin silencing or overexpression induced G2/antephase arrest followed by apoptosis in many but not all cell types. We conclude that pericentrin anchoring of ␥ 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. INTRODUCTIONThe centrosome is the primary microtubule-organizing center in animal cells. At the centrosome core is a pair of barrel-shaped microtubule assemblies, the centrioles (Doxsey, 2001). Centrioles are capable of self-assembly (Marshall et al., 2001;Khodjakov et al., 2002) and can serve as templates for recruitment and organization of the surrounding pericentriolar matrix (Bobinnec et al., 1998;Kirkham et al., 2003). The pericentriolar material or centrosome matrix contains a high proportion of coiled coil proteins and is the site of microtubule nucleation. Within the matrix are large protein complexes of ␥ tubulin and associated proteins that have a ring-like structure and mediate the nucleation of microtubules called ␥ tubulin ring complexes or ␥ TuRCs (Moritz et al., 1995a;Zheng et al., 1995). Other proteins may share the ability to nucleate microtubules because centrosomes can organize microtubules in the absence of functional ␥ tubulin (Sampaio et al., 2001;Strome et al., 2001;Hannak et al., 2002).During cell cycle progression, centrosomes "mature" by recruiting additional ␥ TuRCs and several other proteins, resulting in an increase in the nucleation capacity of the centrosome (reviewed in Blagden and Glover, 2003). However, we still know very little about proteins that directly anchor ␥ TuRCs to centrosomes in vertebrate cells. ...

show abstract

Characterization of Two Related Drosophila γ-tubulin Complexes that Differ in Their Ability to Nucleate Microtubules

Cited by 292 publications

References 45 publications

Early asymmetries in maternal transcript distribution associated with a cortical microtubule network and a polar body in the beetle Tribolium castaneum

Early asymmetries in maternal transcript distribution associated with a cortical microtubule network and a polar body in the beetle Tribolium castaneum

Evolution of the cytoskeleton

Mitosis-specific Anchoring of γ Tubulin Complexes by Pericentrin Controls Spindle Organization and Mitotic Entry

Contact Info

Product

Resources

About