Dissecting protein complexes in branching microtubule nucleation using meiotic xenopus egg extracts

Jae Geun Song, Sabine Petry

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

The mitotic spindle is the microtubule-based apparatus that reliably segregates chromosomes during cell division. Recently, it was discovered that microtubules originate within the mitotic spindle by nucleating off of existing spindle microtubules. This mechanism, termed branching microtubule nucleation, allows the efficient amplification of microtubules while preserving their original polarity as required in the spindle. Three molecular players are known to be involved in this process, namely, the protein TPX2, the protein complex augmin, and the gamma-tubulin ring complex; however, little is known about the assembly of the protein complexes. Here, we use the eight-subunit augmin complex as an example of how to dissect the function and assembly of a protein complex using meiotic Xenopus egg extracts. Specifically, immunodepletion combined with total internal reflection fluorescence (TIRF) microscopy is used to identify the role of the protein complex. In parallel, immunoprecipitation (IP) and tandem mass spectrometry (MS/MS) are used to infer how it is assembled. This approach can be applied to investigate the assembly of other multisubunit protein complexes that function in branching microtubule nucleation and mitotic spindle assembly.

Original languageEnglish (US)
Pages (from-to)728-732
Number of pages5
JournalCold Spring Harbor Protocols
Volume2018
Issue number9
DOIs
StatePublished - Sep 2018

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

Fingerprint Dive into the research topics of 'Dissecting protein complexes in branching microtubule nucleation using meiotic xenopus egg extracts'. Together they form a unique fingerprint.

  • Cite this