Title:
Can we reconstitute a mitotic chromosome from purified components?

Keishi Shintomi
Chromosome Dynamics Laboratory, RIKEN

Abstract:
Chromosome condensation is an essential prerequisite for accurate transmission of genetic information during mitosis. While the unique shape of mitotic chromosomes has attracted attention of numerous cell biologists since the late 19th century, what makes a chromosome and how a chromatin fiber is folded within a chromosome remain enigmatic. To unlock these long-standing questions, I have sought to establish a biochemically tractable system by which a mitotic chromosome is reconstituted from as few purified components as possible in a test tube. Based on our knowledge gained from conventional cell-free systems of frog eggs, I focused on three major chromosomal constituents, namely core histones, type II DNA topoisomerase, and condensin I, and attempted to set up a protocol in which they were properly targeted to genomic DNA in an ordered fashion. Extensive survey allowed me to find a promising condition in which “chromosome-like” structures were assembled from only these three components. Intriguingly, successful assembly of such structures requires removal of tail regions of histone H3-H4, suggesting that physical interaction between nucleosomes might be coordinated during chromosome condensation. Future applications of the reconstitution system to various problems in chromosome biology will be discussed.

References:
1. Shintomi K and Hirano T (2011). The relative ratio of condensin I to II determines chromosome shapes. Genes Dev . 25, 1464-1469.

2. Hirano T (2012). Condensins: universal organizers of chromosomes with diverse functions.Genes Dev . 26, 1659-1678.