Similar to interphase cells, localization of Brd4 to mitotic chromosomes is attributed at least in part to its binding to acetylated histones. In mitotic cells, Brd4 predominantly resides in the long axes of all chromosomes except centromeres where Brd4 is conspicuously absent. An additional feature of Brd4 is that it localizes to chromosomes during mitosis ( Dey et al., 2000). Extending these observations, we found that Brd4 and the related Brd2 recognize acetylated histone codes through the bromodomains ( Kanno et al., 2004) and that acetylated K14 on H3 and K5/12 on H4 are preferred binding sites of Brd4 ( Dey et al., 2003). Previous analysis with fluorescence loss in photobleaching showed that Brd4 dynamically interacts with acetylated histones H3 and H4 in living interphase cells ( Dey et al., 2003). The 200-kDa nuclear protein contains two bromodomains and a long C-terminal domain. Brd4 plays an integral part in a cellular response to drug-induced mitotic stress by preserving a properly acetylated chromatin status.īrd4 is a mammalian bromodomain protein that belongs to the conserved BET family ( Dey et al., 2000). The histone deacetylase inhibitor trichostatin A increased global histone acetylation and perturbed nocodazole-induced Brd4 unloading. The reloading defect observed in Brd4+/- cells coincided with selective hypoacetylation of lysine residues on H3 and H4. Consequently, Brd4+/- cells were impaired in their ability to recover from nocodazole-induced mitotic arrest: a large fraction of +/- cells failed to reach anaphase after drug withdrawal, and those that entered anaphase showed an increased frequency of abnormal chromosomal segregation. However, cells in which a Brd4 allele was disrupted (Brd4+/-), and expressing only half of the normal Brd4 levels, were defective in reloading Brd4 onto chromosomes. Yet, when nocodazole was withdrawn, Brd4 was reloaded onto chromosomes, and cells proceeded to complete cell division. We found that Brd4 was rapidly released from chromosomes upon treatment with antimicrotubule drugs, including the reversible agent nocodazole.
These drugs activate multiple response pathways and arrest cells at mitosis. Mitotic cells are susceptible to antimicrotubule drugs. The mammalian bromodomain protein Brd4 interacts with mitotic chromosomes by binding to acetylated histone H3 and H4 and is thought to play a role in epigenetic memory.
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