Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function

Tien-chen Lin; Linda Gombos; Annett Neuner; Dominik Sebastian; Jesper V Olsen; Ajla Hrle; Christian Benda; Elmar Schiebel

doi:10.1371/journal.pone.0019700

Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function

Tien-chen Lin, Linda Gombos, Annett Neuner, Dominik Sebastian, Jesper V Olsen, Ajla Hrle, Christian Benda, Elmar Schiebel

Novo Nordisk Foundation Center for Protein Research

36 Citations (Scopus)

Abstract

The yeast γ-tubulin Tub4 is assembled with Spc97 and Spc98 into the small Tub4 complex. The Tub4 complex binds via the receptor proteins Spc72 and Spc110 to the spindle pole body (SPB), the functional equivalent of the mammalian centrosome, where the Tub4 complex organizes cytoplasmic and nuclear microtubules. Little is known about the regulation of the Tub4 complex. Here, we isolated the Tub4 complex with the bound receptors from yeast cells. Analysis of the purified Tub4 complex by mass spectrometry identified more than 50 phosphorylation sites in Spc72, Spc97, Spc98, Spc110 and Tub4. To examine the functional relevance of the phosphorylation sites, phospho-mimicking and non-phosphorylatable mutations in Tub4, Spc97 and Spc98 were analyzed. Three phosphorylation sites in Tub4 were found to be critical for Tub4 stability and microtubule organization. One of the sites is highly conserved in γ-tubulins from yeast to human.

Original language	English
Journal	P L o S One
Volume	6
Issue number	5
Pages (from-to)	e19700
ISSN	1932-6203
DOIs	https://doi.org/10.1371/journal.pone.0019700
Publication status	Published - 2011

Keywords

Amino Acid Sequence
Amino Acids
Cell Cycle Proteins
Cytoskeletal Proteins
Humans
Indoleacetic Acids
Microbial Viability
Microtubule-Associated Proteins
Microtubules
Mitotic Spindle Apparatus
Molecular Sequence Data
Mutant Proteins
Nuclear Proteins
Phenotype
Phosphorylation
Protein Binding
Protein Subunits
Protein Transport
Saccharomyces cerevisiae
Saccharomyces cerevisiae Proteins
Tubulin

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10.1371/journal.pone.0019700

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title = "Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function",

abstract = "The yeast γ-tubulin Tub4 is assembled with Spc97 and Spc98 into the small Tub4 complex. The Tub4 complex binds via the receptor proteins Spc72 and Spc110 to the spindle pole body (SPB), the functional equivalent of the mammalian centrosome, where the Tub4 complex organizes cytoplasmic and nuclear microtubules. Little is known about the regulation of the Tub4 complex. Here, we isolated the Tub4 complex with the bound receptors from yeast cells. Analysis of the purified Tub4 complex by mass spectrometry identified more than 50 phosphorylation sites in Spc72, Spc97, Spc98, Spc110 and Tub4. To examine the functional relevance of the phosphorylation sites, phospho-mimicking and non-phosphorylatable mutations in Tub4, Spc97 and Spc98 were analyzed. Three phosphorylation sites in Tub4 were found to be critical for Tub4 stability and microtubule organization. One of the sites is highly conserved in γ-tubulins from yeast to human.",

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author = "Tien-chen Lin and Linda Gombos and Annett Neuner and Dominik Sebastian and Olsen, {Jesper V} and Ajla Hrle and Christian Benda and Elmar Schiebel",

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doi = "10.1371/journal.pone.0019700",

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T1 - Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function

AU - Lin, Tien-chen

AU - Gombos, Linda

AU - Neuner, Annett

AU - Sebastian, Dominik

AU - Olsen, Jesper V

AU - Hrle, Ajla

AU - Benda, Christian

AU - Schiebel, Elmar

PY - 2011

Y1 - 2011

N2 - The yeast γ-tubulin Tub4 is assembled with Spc97 and Spc98 into the small Tub4 complex. The Tub4 complex binds via the receptor proteins Spc72 and Spc110 to the spindle pole body (SPB), the functional equivalent of the mammalian centrosome, where the Tub4 complex organizes cytoplasmic and nuclear microtubules. Little is known about the regulation of the Tub4 complex. Here, we isolated the Tub4 complex with the bound receptors from yeast cells. Analysis of the purified Tub4 complex by mass spectrometry identified more than 50 phosphorylation sites in Spc72, Spc97, Spc98, Spc110 and Tub4. To examine the functional relevance of the phosphorylation sites, phospho-mimicking and non-phosphorylatable mutations in Tub4, Spc97 and Spc98 were analyzed. Three phosphorylation sites in Tub4 were found to be critical for Tub4 stability and microtubule organization. One of the sites is highly conserved in γ-tubulins from yeast to human.

AB - The yeast γ-tubulin Tub4 is assembled with Spc97 and Spc98 into the small Tub4 complex. The Tub4 complex binds via the receptor proteins Spc72 and Spc110 to the spindle pole body (SPB), the functional equivalent of the mammalian centrosome, where the Tub4 complex organizes cytoplasmic and nuclear microtubules. Little is known about the regulation of the Tub4 complex. Here, we isolated the Tub4 complex with the bound receptors from yeast cells. Analysis of the purified Tub4 complex by mass spectrometry identified more than 50 phosphorylation sites in Spc72, Spc97, Spc98, Spc110 and Tub4. To examine the functional relevance of the phosphorylation sites, phospho-mimicking and non-phosphorylatable mutations in Tub4, Spc97 and Spc98 were analyzed. Three phosphorylation sites in Tub4 were found to be critical for Tub4 stability and microtubule organization. One of the sites is highly conserved in γ-tubulins from yeast to human.

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KW - Microtubule-Associated Proteins

KW - Microtubules

KW - Mitotic Spindle Apparatus

KW - Molecular Sequence Data

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KW - Nuclear Proteins

KW - Phenotype

KW - Phosphorylation

KW - Protein Binding

KW - Protein Subunits

KW - Protein Transport

KW - Saccharomyces cerevisiae

KW - Saccharomyces cerevisiae Proteins

KW - Tubulin

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DO - 10.1371/journal.pone.0019700

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SP - e19700

JO - PLoS Computational Biology

JF - PLoS Computational Biology

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ER -

Phosphorylation of the yeast γ-tubulin Tub4 regulates microtubule function

Abstract

Keywords

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