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Free keywords:
Adenosine Triphosphatases/genetics/metabolism
Animals
Cations, Divalent
Cell Line, Tumor
Chickens
Chromatin/drug effects/*metabolism/ultrastructure
Chromatin Assembly and Disassembly/*drug effects
DNA-Binding Proteins/genetics/metabolism
Histones/genetics/*metabolism
Humans
Interphase/drug effects
Lymphocytes/drug effects/*metabolism/ultrastructure
Magnesium/pharmacology
Microscopy, Atomic Force
Mitosis/drug effects
Multiprotein Complexes/genetics/metabolism
Nocodazole/pharmacology
Phosphorylation
*Protein Processing, Post-Translational
Tubulin Modulators/pharmacology
*chromatin
*chromosome compaction
*histone modifications
*mitosis
Abstract:
How eukaryotic chromosomes are compacted during mitosis has been a leading question in cell biology since the nineteenth century. Non-histone proteins such as condensin complexes contribute to chromosome shaping, but appear not to be necessary for mitotic chromatin compaction. Histone modifications are known to affect chromatin structure. As histones undergo major changes in their post-translational modifications during mitotic entry, we speculated that the spectrum of cell-cycle-specific histone modifications might contribute to chromosome compaction during mitosis. To test this hypothesis, we isolated core histones from interphase and mitotic cells and reconstituted chromatin with them. We used mass spectrometry to show that key post-translational modifications remained intact during our isolation procedure. Light, atomic force and transmission electron microscopy analysis showed that chromatin assembled from mitotic histones has a much greater tendency to aggregate than chromatin assembled from interphase histones, even under low magnesium conditions where interphase chromatin remains as separate beads-on-a-string structures. These observations are consistent with the hypothesis that mitotic chromosome formation is a two-stage process with changes in the spectrum of histone post-translational modifications driving mitotic chromatin compaction, while the action of non-histone proteins such as condensin may then shape the condensed chromosomes into their classic mitotic morphology.
