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In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells
To investigate the mechanisms of spindle elongation and chromosome separation in the fission yeast Schizosaccharomyces pombe, we have developed an in vitro assay using a temperature-sensitive mutant strain, nuc2. At the restrictive temperature, nuc2 cells are arrested at a metaphase-like stage with...
Formato: | Texto |
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Lenguaje: | English |
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The Rockefeller University Press
1990
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2116017/ https://www.ncbi.nlm.nih.gov/pubmed/2404993 |
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collection | PubMed |
description | To investigate the mechanisms of spindle elongation and chromosome separation in the fission yeast Schizosaccharomyces pombe, we have developed an in vitro assay using a temperature-sensitive mutant strain, nuc2. At the restrictive temperature, nuc2 cells are arrested at a metaphase-like stage with short spindles and condensed chromosomes. After permeabilization of spheroplasts of the arrested cells, spindle elongation was reactivated by addition of ATP and neurotubulin both at the restrictive and the permissive temperatures, but chromosome separation was not. This suggests that the nuc2 cells are impaired in function at a stage before sister chromatid disjunction. Spindle elongation required both ATP and exogenous tubulin and was inhibited by adenylyl imidodiphosphate (AMPPNP) or vanadate. The ends of yeast half-spindle microtubules pulse-labeled with biotinylated tubulin moved past each other during spindle elongation and a gap formed between the original half-spindles. These results suggest that the primary mechanochemical event responsible for spindle elongation is the sliding apart of antiparallel microtubules of the two half-spindles. |
format | Text |
id | pubmed-2116017 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1990 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21160172008-05-01 In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells J Cell Biol Articles To investigate the mechanisms of spindle elongation and chromosome separation in the fission yeast Schizosaccharomyces pombe, we have developed an in vitro assay using a temperature-sensitive mutant strain, nuc2. At the restrictive temperature, nuc2 cells are arrested at a metaphase-like stage with short spindles and condensed chromosomes. After permeabilization of spheroplasts of the arrested cells, spindle elongation was reactivated by addition of ATP and neurotubulin both at the restrictive and the permissive temperatures, but chromosome separation was not. This suggests that the nuc2 cells are impaired in function at a stage before sister chromatid disjunction. Spindle elongation required both ATP and exogenous tubulin and was inhibited by adenylyl imidodiphosphate (AMPPNP) or vanadate. The ends of yeast half-spindle microtubules pulse-labeled with biotinylated tubulin moved past each other during spindle elongation and a gap formed between the original half-spindles. These results suggest that the primary mechanochemical event responsible for spindle elongation is the sliding apart of antiparallel microtubules of the two half-spindles. The Rockefeller University Press 1990-02-01 /pmc/articles/PMC2116017/ /pubmed/2404993 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 In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title | In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title_full | In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title_fullStr | In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title_full_unstemmed | In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title_short | In vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
title_sort | in vitro reactivation of spindle elongation in fission yeast nuc2 mutant cells |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2116017/ https://www.ncbi.nlm.nih.gov/pubmed/2404993 |