Mitotic chromosomes fold as compact arrays of chromatin loops. To identify the pathway of mitotic chromosome formation, we combined imaging and Hi-C of synchronous DT40 cell cultures with polymer simulations. We show that in prophase, the interphase organization is rapidly lost in a condensin-dependent manner and arrays of consecutive 60 kb loops are formed. During prometaphase ~80 kb inner loops are nested within ~400 kb outer loops. The loop array acquires a helical arrangement with consecutive loops emanating from a central spiral-staircase condensin scaffold. The size of helical turns progressively increases during prometaphase to ~12 Mb. Acute depletion of condensin I or II shows that nested loops form by differential action of the two condensins while condensin II is required for helical winding.
Mitotic chromosome formation involves a relatively minor condensation of the chromatin volume coupled with a dramatic reorganization into the characteristic “X” shape. Here we report results of a detailed morphological analysis, which revealed that chromokinesin KIF4 cooperated in a parallel pathway with condensin complexes to promote the lateral compaction of chromatid arms. In this analysis, KIF4 and condensin were mutually dependent for their dynamic localization on the chromatid axes. Depletion of either caused sister chromatids to expand and compromised the “intrinsic structure” of the chromosomes (defined in an in vitro assay), with loss of condensin showing stronger effects. Simultaneous depletion of KIF4 and condensin caused complete loss of chromosome morphology. In these experiments, topoisomerase IIα contributed to shaping mitotic chromosomes by promoting the shortening of the chromatid axes and apparently acting in opposition to the actions of KIF4 and condensins. These three proteins are major determinants in shaping the characteristic mitotic chromosome morphology.
The 20S cyclosome complex (also known as the anaphase-promoting complex) has ubiquitin ligase activity and is required for mitotic cyclin destruction and sister chromatid separation. The formation and activation of the 20S cyclosome complex is regulated by an unknown mechanism. Here we show that Cut4 (ref. 6) is an essential component of the cyclosome in fission yeast. Cut4 shares sequence similarity with BimE, a protein that regulates mitosis in Aspergillus nidulans. Mutations in cut4 result in hypersensitivity to cyclic AMP and to stress-inducing heavy metals, inhibition of the onset of anaphase, disruption of the 20S complex, and inhibition of mitotic cyclin ubiquitination. These phenotypes are fully suppressed by cAMP phosphodiesterase and the protein kinase A (PKA) regulatory subunit and weakly suppressed by Sti1 (an activator of the Hsp70 and Hsp90 chaperones). Suppression correlates with the amount of 20S complex, indicating that cyclosome formation and activation is inhibited by the cAMP/PKA pathway.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.