RRC ID 53895
Author Shintomi K, Takahashi TS, Hirano T.
Title Reconstitution of mitotic chromatids with a minimum set of purified factors.
Journal Nat Cell Biol
Abstract The assembly of mitotic chromosomes, each composed of a pair of rod-shaped chromatids, is an essential prerequisite for accurate transmission of the genome during cell division. It remains poorly understood, however, how this fundamental process might be achieved and regulated in the cell. Here we report an in vitro system in which mitotic chromatids can be reconstituted by mixing a simple substrate with only six purified factors: core histones, three histone chaperones (nucleoplasmin, Nap1 and FACT), topoisomerase II (topo II) and condensin I. We find that octameric nucleosomes containing the embryonic variant H2A.X-F are highly susceptible to FACT and function as the most productive substrate for subsequent actions of topo II and condensin I. Cdk1 phosphorylation of condensin I is the sole mitosis-specific modification required for chromatid reconstitution. This experimental system will enhance our understanding of the mechanisms of action of individual factors and their cooperation during this process.
Volume 17(8)
Pages 1014-23
Published 2015-8-1
DOI 10.1038/ncb3187
PII ncb3187
PMID 26075356
MeSH Adenosine Triphosphatases / metabolism Animals CDC2 Protein Kinase / metabolism Chromatids / enzymology* Chromatin Assembly and Disassembly* DNA Topoisomerases, Type II / metabolism DNA-Binding Proteins / metabolism HeLa Cells High Mobility Group Proteins / metabolism Histones / genetics Histones / metabolism* Humans Male Mitosis* Molecular Chaperones / genetics Molecular Chaperones / metabolism* Multiprotein Complexes / metabolism Nucleoplasmins / metabolism Nucleosomes / enzymology Phosphorylation Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins / genetics Saccharomyces cerevisiae Proteins / metabolism* Spermatozoa / enzymology* Transcriptional Elongation Factors / metabolism Transfection Xenopus Proteins / genetics Xenopus Proteins / metabolism* Xenopus laevis
IF 20.042
Times Cited 79
DNA material Genome Network Project Human cDNA IRAL008P01 (HGY083561)