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Self-organization of stabilized microtubules by both spindle and midzone mechanisms in Xenopus egg cytosol.
Mitchison, Timothy J; Nguyen, Phuong; Coughlin, Margaret; Groen, Aaron C.
Afiliação
  • Mitchison TJ; Marine Biological Laboratory, Woods Hole, MA 02543, USA.
Mol Biol Cell ; 24(10): 1559-73, 2013 May.
Article em En | MEDLINE | ID: mdl-23515222
ABSTRACT
Previous study of self-organization of Taxol-stabilized microtubules into asters in Xenopus meiotic extracts revealed motor-dependent organizational mechanisms in the spindle. We revisit this approach using clarified cytosol with glycogen added back to supply energy and reducing equivalents. We added probes for NUMA and Aurora B to reveal microtubule polarity. Taxol and dimethyl sulfoxide promote rapid polymerization of microtubules that slowly self-organize into assemblies with a characteristic morphology consisting of paired lines or open circles of parallel bundles. Minus ends align in NUMA-containing foci on the outside, and plus ends in Aurora B-containing foci on the inside. Assemblies have a well-defined width that depends on initial assembly conditions, but microtubules within them have a broad length distribution. Electron microscopy shows that plus-end foci are coated with electron-dense material and resemble similar foci in monopolar midzones in cells. Functional tests show that two key spindle assembly factors, dynein and kinesin-5, act during assembly as they do in spindles, whereas two key midzone assembly factors, Aurora B and Kif4, act as they do in midzones. These data reveal the richness of self-organizing mechanisms that operate on microtubules after they polymerize in meiotic cytoplasm and provide a biochemically tractable system for investigating plus-end organization in midzones.
Assuntos

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Citosol / Microtúbulos / Fuso Acromático Limite: Animals Idioma: En Ano de publicação: 2013 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Citosol / Microtúbulos / Fuso Acromático Limite: Animals Idioma: En Ano de publicação: 2013 Tipo de documento: Article