Shaping Mitotic Chromosomes: From Universal Principles to Local Variations

Abstract

Mitotic chromosomes are often described and considered to be uniform structures that do not vary between different cell types. The models proposed to describe the architecture of mitotic chromosomes reinforce this image, with a uniform series of loops as their foundation. In this thesis, we have challenged certain aspects of these models, without undermining the general concept of spiral-folded arrays of loops. In chapter 2, we describe how the size of mitotic chromosomes shrinks along with cell size during embryo development, by adjusting the amount of condensin I on the chromosome. After fertilization, embryos undergo a series of rapid cell divisions without cell growth, resulting in smaller cells. It has been observed in various organisms that mitotic chromosomes adapt to their cellular environment and thereby become smaller. Our results show that the length compaction of mitotic chromosomes is indeed accompanied by larger loops and a reduced presence of condensin I on the chromosomes. This observation raised an important question: if mitotic chromosomes are universally formed by condensin via stochastic binding, how can cells regulate the size of their chromosomes? To investigate this further, we described in chapter 3 the inverse correlation between the binding of histone H1.8 to mitotic chromosomes and the binding of Top2A and condensin I. Cells can tailor the structure of mitotic chromosomes by regulating the presence of histone H1.8. For example, histone H1.8 showed increased presence on chromosomes after fertilization, consistent with the results from chapter 3. This mechanism enables cells to regulate the size of mitotic chromosomes without strictly controlling the levels of condensin I and II. In the context of chapters 2 and 3, we show in chapter 4 that within a single mitotic chromosome the helicity is not constant. Changes in helical turns correlate with large-scale epigenetic domains from interphase. Our results show that variations in helicity results from an interplay between condensin I and condensin II. While condensin II is essential for helicity, we discovered that condensin I modulates helicity. Although condensin I does not initiate helicity by itself. These results demonstrate that it is the interplay between the machines that fold the chromosome that determines the final organization, rather than each machine functioning independently as current models propose. In chapter 5 we investigated the interaction between condensins and topoisomerase IIα, in particular how the dynamics between these machines influence the topology of mitotic chromosomes. To study topology, we used MC-3C, a 3C technique that examines multiple simultaneous interactions within cells to study local entanglement. Our results show that mitotic chromosomes are highly self-entangled, partly as a result of Top2A activity. However, even in the absence of Top2A, large condensin II loops appear prone to weaving through each other, resulting in a network of interlinked loops. This topological state had not been previously observed and not only shows how loops are intertwined, but also how chromosomes obtain their mechanical properties.