Topoisomerase inhibitor induced dephosphorylation of H1 and H3 histones as a consequence of cell cycle arrest

Abstract

Posttranslational modifications of histones have an integral function in the structural and functional organization of chromatin. Several changes in the modification state of histones could be observed after induction of apoptosis with topoisomerase inhibitors and other inducers. Most of these studies include the analysis of the state of phosphorylation of histones, and the results are to some extent controversial, depending on cell lines and agents used. In the present study we compared the kinetics of the dephosphorylation of H1 and H3 histones with apoptosis markers after treatment of leukemic cell lines with topoisomerase inhibitors. In parallel, we determined cell cycle parameters in detail. Dephosphorylation of both histone classes started within 1 h of induction, and no direct correlation with timing and intensity of the investigated apoptotic features could be observed. In contrast, we show that the effect of topoisomerase inhibitors on the state of H1 and H3 phosphorylation is not directly related to apoptosis, but reflects the changes in the cell cycle distribution of cells treated with these inducers.

Patterns of post-translational histone modifications, chromatin condensation and DNA fragmentation during apoptosis

“…So it might be assumed, that after induction of apoptosis, H3 histones are not subject to an increase in acetylation. However, those changes observed might readily be referred to a strong tendency of dephosphorylation, an observation that has already be described for both H3 at serine 10 and 28 and histone H1 (Happel et al, 2005). As this is still an assumption, it remains to be elucidated which other modifications might be responsible for the changes in peak patterns.…”

“…All these processes will inevitably lead to apoptosis (Caserini et al, 1997). These results were most recently supplemented by Happel et al (Happel et al, 2005), who observed a strong dephosphorylation of histones H3 serine 10 and 28 and histone H1 due to the treatment of different cell types with topotecan ® , which led to a cell cycle arrest.…”

“…Phosphorylation of histone H3 at serines 10 and 28 as well as phosphorylation of H2AX at serine 139 in particular (Rogakou et al, 1998) were first thought to react as direct markers for apoptosis. But these results had to be revised: phosphorylation of histone H3 at serines 10 and 28 has consistently been reported to be associated with mitotic chromatin condensation and cell cycle events (Cheung et al, 2000;Happel et al, 2005;Hendzel et al, 1998b). A direct connection of these modifications with apoptosis could not be confirmed.…”

“…Hendzel et al (Hendzel et al, 1998b) earlier reported that phosphorylation at H3 serine 10 started at G 2 /M transition of the cell cycle and was then initially located to pericentric heterochromatin of each chromosome. By means of flow cytometry and Western blotting, Happel et al (Happel et al, 2005) most recently could provide fundamental evidence that histone H3 phosphorylation at serine 10 and 28 was strongly decreased 1 h after induction of apoptosis in HL-60 cells with either topotecan ® or in HeLa cells with etoposide (both topoisomerase inhibitors). Dephosphorylation of both histone sites S10 and S28 did not correlate with the timing and intensity of the apoptotic parameters, which were measured in parallel.…”

“…As published in recent works, treatment of cells with cytotoxic agents like topotecan ® causes DNA double-strand breaks and a dephosphorylation of histone H3 at serines 10 and 28 (Happel et al, 2005). Moreover, it is widely accepted that transcriptional inactivation and gene silencing correlates with a general histone hypoacetylation of histones H3 and H4 in particular (Grunstein, 1997;Tamburini et al, 2005;Turner, 2000).…”