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Review
. 2021 Sep:117:7-29.
doi: 10.1016/j.semcdb.2021.03.014. Epub 2021 Apr 6.

Mitotic chromosomes

James R Paulson  1 , Damien F Hudson  2 , Fernanda Cisneros-Soberanis  3 , William C Earnshaw  4
Affiliations
Review

Mitotic chromosomes

James R Paulson et al. Semin Cell Dev Biol. 2021 Sep.

Abstract

Our understanding of the structure and function of mitotic chromosomes has come a long way since these iconic objects were first recognized more than 140 years ago, though many details remain to be elucidated. In this chapter, we start with the early history of chromosome studies and then describe the path that led to our current understanding of the formation and structure of mitotic chromosomes. We also discuss some of the remaining questions. It is now well established that each mitotic chromatid consists of a central organizing region containing a so-called "chromosome scaffold" from which loops of DNA project radially. Only a few key non-histone proteins and protein complexes are required to form the chromosome: topoisomerase IIα, cohesin, condensin I and condensin II, and the chromokinesin KIF4A. These proteins are concentrated along the axis of the chromatid. Condensins I and II are primarily responsible for shaping the chromosome and the scaffold, and they produce the loops of DNA by an ATP-dependent process known as loop extrusion. Modelling of Hi-C data suggests that condensin II adopts a spiral staircase arrangement with an extruded loop extending out from each step in a roughly helical pattern. Condensin I then forms loops nested within these larger condensin II loops, thereby giving rise to the final compaction of the mitotic chromosome in a process that requires Topo IIα.

Keywords: Chromosome; Cohesin; Condensin; KIF4; Scaffold; Topoisomerase IIα.

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Figures

Fig. 1
Fig. 1
Selective timeline of key events in mitotic chromosome studies. For simplicity, references are omitted from the figure, but these are given in the relevant sections of the text.
Fig. 2
Fig. 2
Isolation and characterization of the mitotic chromosome scaffold fraction. (A) General flow-chart for the isolation of mitotic chromosome scaffolds. (B) SDS polyacrylamide gel of mitotic chromosomes (lane 1) and the scaffold fraction isolated by treatment with nucleases and 2 M NaCl (lane 2) from chicken MSB-1 cells . (C) Human (HeLa) mitotic chromosome scaffold isolated at low ionic strength with dextran sulphate/heparin and centrifuged onto an electron microscope grid .
Fig. 3
Fig. 3
Human mitotic chromosome showing radial loops with nucleosomes. (A) Chromosomes were isolated from HeLa cells, expanded at low ionic strength in the absence of divalent cations using TEEN buffer, centrifuged onto an electron microscope grid and then treated with aqueous uranyl acetate (which caused the chromatin not adherent to the carbon film to collapse back onto the chromatid axes) . (B) Enlarged view of the boxed area. Radial loops with abundant nucleosomes are clearly seen.
Fig. 4
Fig. 4
Scaffold region of a histone-depleted human mitotic chromosome. Chromosomes isolated from HeLa cells were depleted of histones by treatment with 2 M NaCl and prepared by Kleinschmidt spreading . Note the open, network-like structure of the scaffold in the chromatid on the left. The dense “core” in the chromatid on the right and the “straps” emanating from it are most likely due to collapse during dehydration and staining of DNA that was not completely adsorbed to the cytochrome c monolayer.
Fig. 5
Fig. 5
Localization of condensin and KIF4A to the chromatid axis in mitotic cells. (A,B) Two mitotic DT40 cells with conditional SMC2 knockout (SMC2OFF) stained for endogenous KIF4A (red) and expressing SMC2-TrAP (green - stained with anti-SBP antibody). Both proteins are localized to the axis of sister chromatids (DNA - blue). Unpublished images provided by Kumiko Samejima from .
Fig. 6
Fig. 6
3D-Reconstruction of human metaphase chromosomes obtained by SBF-SEM. (A) Section 84 of 225 of a metaphase hTERT RPE1 cell imaged by Serial Block-Face Scanning Electron Microscopy (SBF-SEM) using 4 nm resolution in XY and 75 nm in Z. The microscope was a Gatan 3View. (B) Modelling and 3D reconstruction of chromosomes. Representative images in XY and ZY are shown. (C) Segmentation and labelling of chromosomes from the 3D reconstruction. 46 chromosomes were resolved and each is shown in a different colour. Chromosomes 1–5 and 19–22 are identified unambiguously by their relative lengths and volumes. Image processing used the program AMIRA (Thermo Scientific).
Fig. 7
Fig. 7
Organization and protein composition of the three SMC protein complexes. Nomenclature is for the vertebrate proteins. For a description of the nomenclature in various model organisms, see .
Fig. 8
Fig. 8
Structural dynamics of SMC complexes. (A) ATP binding regulates the pairing of SMC protein heads. The heads pair, closing the SMC ring, when ATP is bound, and open following ATP hydrolysis. The strap-like kleisin subunit which also joins the two heads together, is not shown in this diagram. (B) The coiled-coils of SMC proteins are highly dynamic. They can form either a ring , , a closed I-structure , , , , , a folded-over I-structure , or a butterfly structure . The distribution of these different states appears to be regulated by ATP-binding , . Since both the hinge region and the HEAT repeat subunits can bind DNA, it is highly likely that these conformational changes contribute to loop extrusion, although the detailed mechanism is still under investigation.
Fig. 9
Fig. 9
Condensin is required to establish a stable mitotic chromosome architecture. Chromosomes in mitotic chicken DT40 cells either containing SMC2 (A) or depleted of SMC2 and therefore condensin (B) were placed in low ionic strength TEEN buffer in the absence of divalent cations . (A) Chromosomes in cells containing SMC2 expand, but retain a recognizable chromosome morphology. (B) Chromosomes in cells depleted of condensin look relatively normal initially though slightly swollen (left-hand micrograph), but after addition of TEEN buffer, they unravel completely. Experiment performed by Paola Vagnarelli .
Fig. 10
Fig. 10
Nested loop extrusion by Condensin I and II. (A) Condensin II begins loop extrusion during prophase, so that by late prophase the average loop size is about 60 kb. By metaphase, the loop size has grown to around 400 kb, but the observed loops are smaller because after nuclear envelope breakdown (NEB) condensin I jumps onto the DNA within the condensin II loops and begins to extrude its own loops . This process depends on the fact that the association of condensin II with DNA is substantially more stable than that of condensin I . (B) Frames from an animation describing mitotic chromosome organization made by Anton Goloborodko .
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