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Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin
A trimeric complex formed by Tub4p, the budding yeast γ-tubulin, and the two spindle pole body components, Spc98p and Spc97p, has recently been characterized in Saccharomyces cerevisiae. We reasoned that crucial functions, such as the control of microtubule nucleation, could be maintained among dive...
Autores principales: | , , , |
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Formato: | Texto |
Lenguaje: | English |
Publicado: |
The Rockefeller University Press
1998
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2132749/ https://www.ncbi.nlm.nih.gov/pubmed/9566969 |
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author | Tassin, Anne-Marie Celati, Claude Moudjou, Mohammed Bornens, Michel |
author_facet | Tassin, Anne-Marie Celati, Claude Moudjou, Mohammed Bornens, Michel |
author_sort | Tassin, Anne-Marie |
collection | PubMed |
description | A trimeric complex formed by Tub4p, the budding yeast γ-tubulin, and the two spindle pole body components, Spc98p and Spc97p, has recently been characterized in Saccharomyces cerevisiae. We reasoned that crucial functions, such as the control of microtubule nucleation, could be maintained among divergent species. SPC98-related sequences were searched in dbEST using the BLASTN program. Primers derived from the human expressed sequence tag matching SPC98 were used to clone the 5′ and 3′ cDNA ends by rapid amplification of cDNA ends (RACE)-PCR. The human Spc98 cDNA presents an alternative splicing at the 3′ end. The deduced protein possesses 22% identity and 45% similarity with the yeast homologue. We further report that the human Spc98p, like γ-tubulin, is concentrated at the centrosome, although a large fraction is found in cytosolic complexes. Sucrose gradient sedimentation of the cytosolic fraction and immunoprecipitation experiments demonstrate that both γ-tubulin and HsSpc98p are in the same complex. Interestingly, Xenopus sperm centrosomes, which are incompetent for microtubule nucleation before their activation in the egg cytoplasm, were found to contain similar amounts of both Spc98p and γ-tubulin to human somatic centrosomes, which are competent for microtubule nucleation. Finally, affinity-purified antibodies against Spc98p inhibit microtubule nucleation on isolated centrosomes, as well as in microinjected cells, suggesting that this novel protein is indeed required for the nucleation reaction. |
format | Text |
id | pubmed-2132749 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1998 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21327492008-05-01 Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin Tassin, Anne-Marie Celati, Claude Moudjou, Mohammed Bornens, Michel J Cell Biol Articles A trimeric complex formed by Tub4p, the budding yeast γ-tubulin, and the two spindle pole body components, Spc98p and Spc97p, has recently been characterized in Saccharomyces cerevisiae. We reasoned that crucial functions, such as the control of microtubule nucleation, could be maintained among divergent species. SPC98-related sequences were searched in dbEST using the BLASTN program. Primers derived from the human expressed sequence tag matching SPC98 were used to clone the 5′ and 3′ cDNA ends by rapid amplification of cDNA ends (RACE)-PCR. The human Spc98 cDNA presents an alternative splicing at the 3′ end. The deduced protein possesses 22% identity and 45% similarity with the yeast homologue. We further report that the human Spc98p, like γ-tubulin, is concentrated at the centrosome, although a large fraction is found in cytosolic complexes. Sucrose gradient sedimentation of the cytosolic fraction and immunoprecipitation experiments demonstrate that both γ-tubulin and HsSpc98p are in the same complex. Interestingly, Xenopus sperm centrosomes, which are incompetent for microtubule nucleation before their activation in the egg cytoplasm, were found to contain similar amounts of both Spc98p and γ-tubulin to human somatic centrosomes, which are competent for microtubule nucleation. Finally, affinity-purified antibodies against Spc98p inhibit microtubule nucleation on isolated centrosomes, as well as in microinjected cells, suggesting that this novel protein is indeed required for the nucleation reaction. The Rockefeller University Press 1998-05-04 /pmc/articles/PMC2132749/ /pubmed/9566969 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/). |
spellingShingle | Articles Tassin, Anne-Marie Celati, Claude Moudjou, Mohammed Bornens, Michel Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title | Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title_full | Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title_fullStr | Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title_full_unstemmed | Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title_short | Characterization of the Human Homologue of the Yeast Spc98p and Its Association with γ-Tubulin |
title_sort | characterization of the human homologue of the yeast spc98p and its association with γ-tubulin |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2132749/ https://www.ncbi.nlm.nih.gov/pubmed/9566969 |
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