Massive histone H3 phosphorylation without mitotic chromosome condensation

Histone post-translational modifications change dramatically as cells enter mitosis. Indeed, histone phosphorylation and deacetylation have been proposed to be major factors driving the compaction of mitotic chromosomes. However, whether these modification changes directly regulate mitotic chromatin compaction remains a subject of debate. Here, we use a CDK1^as chemical-genetic system to accumulate cells in G₂ phase. Treatment of these G₂ cells with the phosphatase inhibitor calyculin A efficiently causes them to undergo premature chromosome condensation, even when the CDK1^as allele is inhibited with 1NM-PP1. Surprisingly, simultaneous treatment of the cells with R547, an inhibitor of CDK1, 2, and 4 kinases, causes them to remain phenotypically in G₂ phase. Strikingly, although mitotic-like chromatin condensation fails to occur in these cells, LC-MS reveals that histone H3 phosphorylation occurs even more efficiently than in a normal mitosis. Furthermore, histone deacetylation also occurs in those interphase-looking calyculin A-treated cells. These results reveal that histone phosphorylation and deacetylation are not sufficient to drive mitotic-like chromosome condensation and compaction.