Xgrip109: A γ Tubulin–Associated Protein with an Essential Role in γ Tubulin Ring Complex (γTuRC) Assembly and Centrosome Function

Martin, Ona C.; Gunawardane, Ruwanthi N.; Iwamatsu, Akihiro; Zheng, Yixian

doi:10.1083/jcb.141.3.675

Cited by 106 publications

(99 citation statements)

References 42 publications

(78 reference statements)

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“…XLK-WG cells (ATCC, Manassas, Virginia; (Martin et al, 1998) were cultured as directed by the supplier: 29 C with 5% CO 2 in RPMI-1640 medium (ThermoFisher Scientific, Waltham, MA) with 20% FBS (Invitrogen, Carlsbad, California; Iwamoto et al, 2012;Laub et al, 2010).…”

Section: Methodsmentioning

confidence: 99%

“…However, some gene duplicates show evidence of functional divergence (Chain and Evans, 2006;Hellsten et al, 2007). The objective of this study was to test the hypothesis that endogenous AHR1a and AHR1b have different molecular functions in XLK-WG (Martin et al, 1998), a frog cell line used previously for AHR characterization (Iwamoto et al, 2012;Laub et al, 2010). Using TALENs, we generated mutant cells lacking either AHR1a or AHR1b, examining the responsiveness of each to a xenobiotic and a candidate endogenous agonist.…”

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confidence: 99%

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Subfunctionalization of Paralogous Aryl Hydrocarbon Receptors from the FrogXenopus Laevis: Distinct Target Genes and Differential Responses to Specific Agonists in a Single Cell Type

2016

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Gene duplication confers genetic redundancy that can facilitate subfunctionalization, the partitioning of ancestral functions between paralogs. We capitalize on a recent genome duplication in Xenopus laevis (African clawed frog) to interrogate possible functional differentiation between alloalleles of the aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor that mediates toxicity of dioxin-like compounds and plays a role in the physiology and development of the cardiovascular, hepatic, and immune systems in vertebrates. X. laevis has 2 AHR genes, AHR1a and AHR1b. To test the hypothesis that the encoded proteins exhibit different molecular functions, we used TALENs in XLK-WG cells, generating mutant lines lacking functional versions of each AHR and measuring the transcriptional responsiveness of several target genes to the toxic xenobiotic 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and the candidate endogenous ligand 6-formylindolo[3,2-b]carbazole (FICZ). Mutation of either AHR1a or AHR1b reduced TCDD induction of the canonical AHR target, Cytochrome P4501A6, by 75%, despite the much lower abundance of AHR1b in wild-type cells. More modestly induced target genes, encoding aryl hydrocarbon receptor repressor (AHRR), spectrin repeat-containing nuclear envelope protein 1 (SYNE-1), and gap junction protein gamma 1 (GJC1), were regulated solely by AHR1a. AHR1b was responsible for CYP1A6 induction by FICZ, while AHR1a mediated FICZ induction of AHRR. We conclude that AHR1a and AHR1b have distinct transcriptional functions in response to specific agonists, even within a single cell type. Functional analysis of frog AHR paralogs advances the understanding of AHR evolution and as well as the use of frog models of developmental toxicology such as FETAX.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Subfunctionalization of Paralogous Aryl Hydrocarbon Receptors from the FrogXenopus Laevis: Distinct Target Genes and Differential Responses to Specific Agonists in a Single Cell Type

2016

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“…␥TuRCs from these organisms share a similar structure and protein composition, and they can nucleate MTs in vitro (Zheng et al, 1995;Oegema et al, 1999;Murphy et al, 2001). Additional biochemical and structural studies reveal that ␥TuRC is recruited to the centrosome to mediate MT nucleation (Moritz et al, 1995a(Moritz et al, , 1995bMartin et al, 1998;Schnackenberg et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

Characterization of a New γTuRC Subunit with WD Repeats

Gunawardane

Martin

Zheng

2003

MBoC

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The ␥-tubulin ring complex (␥TuRC), consisting of multiple protein subunits, can nucleate microtubule assembly. Although many subunits of the ␥TuRC have been identified, a complete set remains to be defined in any organism. In addition, how the subunits interact with each other to assemble into ␥TuRC remains largely unknown. Here, we report the characterization of a novel ␥TuRC subunit, Drosophila gamma ring protein with WD repeats (Dgp71WD). With the exception of ␥-tubulin, Dgp71WD is the only ␥TuRC component identified to date that does not contain the grip motifs, which are signature sequences conserved in ␥TuRC components. By performing immunoprecipitations after pair-wise coexpression in Sf9 cells, we show that Dgp71WD directly interacts with the grip motif-containing ␥TuRC subunits, Dgrips84, 91, 128, and 163, suggesting that Dgp71WD may play a scaffolding role in ␥TuRC organization. We also show that Dgrips128 and 163, like Dgrips84 and 91, can interact directly with ␥-tubulin. Coexpression of any of these grip motif-containing proteins with ␥-tubulin promotes ␥-tubulin binding to guanine nucleotide. In contrast, in the same assay Dgp71WD interacts with ␥-tubulin but does not facilitate nucleotide binding.

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“…␥ 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

270

258

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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. ...

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Xgrip109: A γ Tubulin–Associated Protein with an Essential Role in γ Tubulin Ring Complex (γTuRC) Assembly and Centrosome Function

Cited by 106 publications

References 42 publications

Subfunctionalization of Paralogous Aryl Hydrocarbon Receptors from the FrogXenopus Laevis: Distinct Target Genes and Differential Responses to Specific Agonists in a Single Cell Type

Subfunctionalization of Paralogous Aryl Hydrocarbon Receptors from the FrogXenopus Laevis: Distinct Target Genes and Differential Responses to Specific Agonists in a Single Cell Type

Characterization of a New γTuRC Subunit with WD Repeats

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

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