Mitosis-specific phosphorylation of histone H3 initiates primarily within pericentromeric heterochromatin during G2 and spreads in an ordered fashion coincident with mitotic chromosome condensation

Abstract

We have generated and characterized a novel site-specific antibody highly specific for the phosphorylated form of the amino-terminus of histone H3 (Ser10). In this study, we used this antibody to examine in detail the relationship between H3 phosphorylation and mitotic chromosome condensation in mammalian cells. Our results extend previous biochemical studies by demonstrating that mitotic phosphorylation of H3 initiates nonrandomly in pericentromeric heterochromatin in late G2 interphase cells. Following initiation, H3 phosphorylation appears to spread throughout the condensing chromatin and is complete in most cell lines just prior to the formation of prophase chromosomes, in which a phosphorylated, but nonmitotic, chromosomal organization is observed. In general, there is a precise spatial and temporal correlation between H3 phosphorylation and initial stages of chromatin condensation. Dephosphorylation of H3 begins in anaphase and is complete immediately prior to detectable chromosome decondensation in telophase cells. We propose that the singular phosphorylation of the amino-terminus of histone H3 may be involved in facilitating two key functions during mitosis: (1) regulate protein-protein interactions to promote binding of trans-acting factors that "drive" chromatin condensation as cells enter M-phase and (2) coordinate chromatin decondensation associated with M-phase.

The dNTP triphosphohydrolase activity of SAMHD1 persists during S-phase when the enzyme is phosphorylated at T592

“…Securin and cyclin B levels were monitored as indicators of APC/C cdc20 activity, phospho-histone H3 (Ser10) (pH3S10) was monitored as a marker of condensed mitotic chromosomes, and we followed the dephosphorylation of CDK substrates with anti phospho-threonine (pThr-CDK) and anti phospho-serine (pSer-CDK) antibodies. After RO3306 addition, securin and cyclin B remained stable due to the presence of the proteasome inhibitor, while the disappearance of pH3S10 marked the entrance into anaphase [29], (Figure 6(c)). Significant dephosphorylation of pThrCDK sites occurred within 15 min after RO3306 addition, whereas dephosphorylation of pSerCDK sites was slower, reflecting differential dephosphorylation patterns during mitotic exit [27].…”

Chromosomes with delayed replication timing lead to checkpoint activation, delayed recruitment of Aurora B and chromosome instability

“…mitosis-specific phosphorylation of histone H3 on serine 10 ( Figure 1a and b and see Smith et al, 2001). Because H3 phosphorylation is typically initiated in late G 2 and completed just before nuclear envelope breakdown (Hendzel et al, 1997), these observations suggest that the more extreme undercondensed chromosomes retain an 'interphase state' of condensation in cells that contain fully phosphorylated and condensed metaphase chromosomes. This observation, and the 2-3 h DRT (Figure 2a and see Smith et al, 2001), suggested the possibility of active DNA synthesis occurring on these chromosomes during mitosis.…”

Proper expression of the Gcn5 histone acetyltransferase is required for neural tube closure in mouse embryos

“…Phosphorylation of histone H3 at S10 serves as a mitotic marker (Hendzel et al, 1997), and whole mount immunohistochemistry with an antiphospho S10 histone H3 antibody revealed no obvious depletion of mitotic cells in Gcn5 flox(neo)/⌬ mutants at E8.5, E9.5, and E10.5 (data not shown). The neural tube defects in these mice, then, is not likely due to defects in cell proliferation.…”