Abstract
SummaryRecent studies have revealed the importance of Ki-67 and the chromosome periphery in chromosome structure and segregation, but little is known about this elusive chromosome compartment. Here we used correlative light and serial block-face scanning electron microscopy, which we term 3D-CLEM, to model the entire mitotic chromosome complement at ultra-structural resolution. Prophase chromosomes exhibit a highly irregular surface appearance with a volume smaller than metaphase chromosomes. This may be because of the absence of the periphery, which associates with chromosomes only after nucleolar disassembly later in prophase. Indeed, the nucleolar volume almost entirely accounts for the extra volume found in metaphase chromosomes. Analysis of wild-type and Ki-67-depleted chromosomes reveals that the periphery comprises 30%–47% of the entire chromosome volume and more than 33% of the protein mass of isolated mitotic chromosomes determined by quantitative proteomics. Thus, chromatin makes up a surprisingly small percentage of the total mass of metaphase chromosomes.
Highlights
Since their first discovery in 1882 (Flemming, 1882), mitotic chromosomes have been a subject of intense study
We have developed a pipeline for the three-dimensional analysis of chromosomes by correlative light and serial block-face scanning electron microscopy
As a first test of the 3D-CLEM method, we performed a targeted analysis of a synthetic human artificial chromosome (HAC) (Figure 2)
Summary
Oscar Molina, ..., Vladimir Larionov, Ian A. Booth et al develop a microscopy method called 3D-CLEM. Using this approach, they find that a large portion of mitotic chromosomes is not composed of chromatin, challenging classical perceptions of chromosomes. 2016, Molecular Cell 64, 790–802 November 17, 2016 a 2016 The Author(s).
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