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Chromosome segregation and cell division are essential, highly ordered processes that depend on numerous protein complexes. Results from recent RNA interference screens indicate that the identity and composition of these protein complexes is incompletely understood. Using gene tagging on bacterial artificial chromosomes, protein localization, and tandem-affinity purification-mass spectrometry, the MitoCheck consortium has analyzed about 100 human protein complexes, many of which had not or had only incompletely been characterized. This work has led to the discovery of previously unknown, evolutionarily conserved subunits of the anaphase-promoting complex and the gamma-tubulin ring complex--large complexes that are essential for spindle assembly and chromosome segregation. The approaches we describe here are generally applicable to high-throughput follow-up analyses of phenotypic screens in mammalian cells.

Original publication

DOI

10.1126/science.1181348

Type

Journal article

Journal

Science

Publication Date

30/04/2010

Volume

328

Pages

593 - 599

Keywords

Anaphase-Promoting Complex-Cyclosome, Centrosome, Chromosome Segregation, Chromosomes, Artificial, Bacterial, Databases, Genetic, Genomics, Green Fluorescent Proteins, HeLa Cells, Humans, Mitosis, Multiprotein Complexes, Open Reading Frames, Protein Binding, Protein Interaction Mapping, Protein Subunits, RNA Interference, Spindle Apparatus, Tubulin, Ubiquitin-Protein Ligase Complexes