Identification of a Novel Phosphorylation Site on Histone H3 Coupled with Mitotic Chromosome Condensation

Histone H3 (H3) phosphorylation at Ser10 occurs during mitosis in eukaryotes and was recently shown to play an important role in chromosome condensation in Tetrahymena. When producing monoclonal antibodies that recognize glial fibrillary acidic protein phosphorylation at Thr7, we obtained some monoc...

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Bibliographic Details
Published in:The Journal of biological chemistry 1999-09, Vol.274 (36), p.25543-25549
Main Authors: Goto, Hidemasa, Kosako, Hidetaka, Tomono, Yasuko, Ajiro, Kozo, Fujita, Masatoshi, Sakurai, Minoru, Okawa, Katsuya, Iwamatsu, Akihiro, Okigaki, Tohru, Takahashi, Toshitada, Inagaki, Masaki
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Chromosomes - metabolism
Chromosomes - ultrastructure
DNA Mutational Analysis
Histones - genetics
Histones - metabolism
Mitosis
Molecular Sequence Data
Phosphorylation
Rats
Tetrahymena
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Summary:Histone H3 (H3) phosphorylation at Ser10 occurs during mitosis in eukaryotes and was recently shown to play an important role in chromosome condensation in Tetrahymena. When producing monoclonal antibodies that recognize glial fibrillary acidic protein phosphorylation at Thr7, we obtained some monoclonal antibodies that cross-reacted with early mitotic chromosomes. They reacted with 15-kDa phosphoprotein specifically in mitotic cell lysate. With microsequencing, this phosphoprotein was proved to be H3. Mutational analysis revealed that they recognized H3 Ser28phosphorylation. Then we produced a monoclonal antibody, HTA28, using a phosphopeptide corresponding to phosphorylated H3 Ser28. This antibody specifically recognized the phosphorylation of H3 Ser28 but not that of glial fibrillary acidic protein Thr7. Immunocytochemical studies with HTA28 revealed that Ser28 phosphorylation occurred in chromosomes predominantly during early mitosis and coincided with the initiation of mitotic chromosome condensation. Biochemical analyses using32P-labeled mitotic cells also confirmed that H3 is phosphorylated at Ser28 during early mitosis. In addition, we found that H3 is phosphorylated at Ser28 as well as Ser10 when premature chromosome condensation was induced in tsBN2 cells. These observations suggest that H3 phosphorylation at Ser28, together with Ser10, is a conserved event and is likely to be involved in mitotic chromosome condensation.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.274.36.25543