Limits...
Megabase chromatin domains involved in DNA double-strand breaks in vivo.

Rogakou EP, Boon C, Redon C, Bonner WM - J. Cell Biol. (1999)

Bottom Line: When DNA double-strand breaks are introduced into specific partial nuclear volumes of cells by means of a pulsed microbeam laser, gamma-H2AX foci form at these sites.These results offer direct visual confirmation that gamma-H2AX forms en masse at chromosomal sites of DNA double-strand breaks.The results further suggest the possible existence of units of higher order chromatin structure involved in monitoring DNA integrity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Pharmacology, Division of Basic Sciences, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The loss of chromosomal integrity from DNA double-strand breaks introduced into mammalian cells by ionizing radiation results in the specific phosphorylation of histone H2AX on serine residue 139, yielding a specific modified form named gamma-H2AX. An antibody prepared to the unique region of human gamma-H2AX shows that H2AX homologues are phosphorylated not only in irradiated mammalian cells but also in irradiated cells from other species, including Xenopus laevis, Drosophila melanogaster, and Saccharomyces cerevisiae. The antibody reveals that gamma-H2AX appears as discrete nuclear foci within 1 min after exposure of cells to ionizing radiation. The numbers of these foci are comparable to the numbers of induced DNA double-strand breaks. When DNA double-strand breaks are introduced into specific partial nuclear volumes of cells by means of a pulsed microbeam laser, gamma-H2AX foci form at these sites. In mitotic cells from cultures exposed to nonlethal amounts of ionizing radiation, gamma-H2AX foci form band-like structures on chromosome arms and on the end of broken arms. These results offer direct visual confirmation that gamma-H2AX forms en masse at chromosomal sites of DNA double-strand breaks. The results further suggest the possible existence of units of higher order chromatin structure involved in monitoring DNA integrity.

Show MeSH

Related in: MedlinePlus

Interphase and mitotic cells from human MCF7 and M. muntjak cultures. (A–C) MCF7 cells from an unirradiated culture (A) or from cultures exposed to 0.6 Gy and allowed to recover for 15 (B) or 225 (C) min before fixation. (D) M. muntjak cells from a culture exposed to 0.6 Gy and allowed to recover for 15 min before fixation.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2169482&req=5

Figure 5: Interphase and mitotic cells from human MCF7 and M. muntjak cultures. (A–C) MCF7 cells from an unirradiated culture (A) or from cultures exposed to 0.6 Gy and allowed to recover for 15 (B) or 225 (C) min before fixation. (D) M. muntjak cells from a culture exposed to 0.6 Gy and allowed to recover for 15 min before fixation.

Mentions: Mitotic MCF7 cells were present in some of the cultures analyzed in Fig. 2. The mitotic cell noted in Fig. 2 J (m) is of particular interest because this culture was fixed only 3 min after irradiation, a result indicating that γ-H2AX foci form on mitotic chromosomes as well as on interphase chromatin. When fields of cells from MCF7 cultures containing both mitotic and interphase cells were analyzed for γ-H2AX foci 15 and 225 min after exposure to 0.6 Gy, the number of foci was decreased at the latter time in both interphase and mitotic cells (Fig. 5, A–C). These results show that the kinetics of γ-H2AX foci appearance and disappearance are similar whether cells are interphase or mitotic.


Megabase chromatin domains involved in DNA double-strand breaks in vivo.

Rogakou EP, Boon C, Redon C, Bonner WM - J. Cell Biol. (1999)

Interphase and mitotic cells from human MCF7 and M. muntjak cultures. (A–C) MCF7 cells from an unirradiated culture (A) or from cultures exposed to 0.6 Gy and allowed to recover for 15 (B) or 225 (C) min before fixation. (D) M. muntjak cells from a culture exposed to 0.6 Gy and allowed to recover for 15 min before fixation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169482&req=5

Figure 5: Interphase and mitotic cells from human MCF7 and M. muntjak cultures. (A–C) MCF7 cells from an unirradiated culture (A) or from cultures exposed to 0.6 Gy and allowed to recover for 15 (B) or 225 (C) min before fixation. (D) M. muntjak cells from a culture exposed to 0.6 Gy and allowed to recover for 15 min before fixation.
Mentions: Mitotic MCF7 cells were present in some of the cultures analyzed in Fig. 2. The mitotic cell noted in Fig. 2 J (m) is of particular interest because this culture was fixed only 3 min after irradiation, a result indicating that γ-H2AX foci form on mitotic chromosomes as well as on interphase chromatin. When fields of cells from MCF7 cultures containing both mitotic and interphase cells were analyzed for γ-H2AX foci 15 and 225 min after exposure to 0.6 Gy, the number of foci was decreased at the latter time in both interphase and mitotic cells (Fig. 5, A–C). These results show that the kinetics of γ-H2AX foci appearance and disappearance are similar whether cells are interphase or mitotic.

Bottom Line: When DNA double-strand breaks are introduced into specific partial nuclear volumes of cells by means of a pulsed microbeam laser, gamma-H2AX foci form at these sites.These results offer direct visual confirmation that gamma-H2AX forms en masse at chromosomal sites of DNA double-strand breaks.The results further suggest the possible existence of units of higher order chromatin structure involved in monitoring DNA integrity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Pharmacology, Division of Basic Sciences, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The loss of chromosomal integrity from DNA double-strand breaks introduced into mammalian cells by ionizing radiation results in the specific phosphorylation of histone H2AX on serine residue 139, yielding a specific modified form named gamma-H2AX. An antibody prepared to the unique region of human gamma-H2AX shows that H2AX homologues are phosphorylated not only in irradiated mammalian cells but also in irradiated cells from other species, including Xenopus laevis, Drosophila melanogaster, and Saccharomyces cerevisiae. The antibody reveals that gamma-H2AX appears as discrete nuclear foci within 1 min after exposure of cells to ionizing radiation. The numbers of these foci are comparable to the numbers of induced DNA double-strand breaks. When DNA double-strand breaks are introduced into specific partial nuclear volumes of cells by means of a pulsed microbeam laser, gamma-H2AX foci form at these sites. In mitotic cells from cultures exposed to nonlethal amounts of ionizing radiation, gamma-H2AX foci form band-like structures on chromosome arms and on the end of broken arms. These results offer direct visual confirmation that gamma-H2AX forms en masse at chromosomal sites of DNA double-strand breaks. The results further suggest the possible existence of units of higher order chromatin structure involved in monitoring DNA integrity.

Show MeSH
Related in: MedlinePlus