title: Dynamics of radiation induced γH2AX foci in chromatin subcompartments of mouse pachytene spermatocytes and round spermatids.
abstract: Chromatin compaction is thought to influence the severity of radiation-induced DNA damage. We assessed how chromatin state affects DNA double-strand break repair within eu-/heterochromatin domains in male germ cells by profiling the spatiotemporal dynamics of γ-radiation-induced γH2AX foci in confocal images of mouse pachytene spermatocytes and round spermatids (5 min to 16 hr post-irradiation, in vivo). In unirradiated cells, all DNA-dense heterochromatin domains showed compaction by anti-H3K9me3-staining, except for peripheral areas. Following irradiation, this signal was lost within 5 min, but regained later (8-16 hr); these two events coincided with the appearance and loss of γH2AX foci, respectively. While euchromatin showed a large number of bright foci in both cell types, heterochromatin had few foci. In spermatids, a few small, faint foci appeared within chromocenters. Pachytene-stage, on the other hand, lacked foci within heterochromatin, although a few were closely associated with the heterochromatin periphery. The number of euchromatin foci in spermatids showed a dose-dependent enhancement following irradiation (0.5-4 Gy), although no significant increase was seen in the quantity of heterochromatin foci. While all foci in pachytene-stage cells were resolved, spermatids showed large residual foci-especially from heterochromatin foci, which remained faint for up to 4 hr, then increased in size between 8-16 hr, expanding at the chromocenter periphery and eventually protruding into euchromatin at H3K9me3-signal-free areas. Thus, this study identified scant foci formation and poor repair within heterochromatin, with distinctly different dynamics in meiotic and post-meiotic stages of spermatogenesis, and provides direct evidence for heterochromatin decompaction following DNA damage, which facilitates repair/repositioning of foci towards euchromatin domains. It is the first demonstration of spatiotemporal mobilization of double-strand breaks with respect to chromatin subdomains in male germ cells.
authors.affiliation: Cytogenetics Laboratory, Department of Zoology, Centre of Advanced Study, Banaras Hindu University, Varanasi, India.
authors.forename: Priti
authors.initials: P
authors.lastname: Singh
authors.forename: Mercy J
authors.initials: MJ
authors.lastname: Raman
mesh_terms: D000818:Animals; D004249:DNA Damage; D004307:Dose-Response Relationship, Radiation; D005720:Gamma Rays; D006570:Heterochromatin / Q000378:metabolism* / Q000473:pathology; D006657:Histones / Q000378:metabolism*; D008297:Male; D051379:Mice; D049471:Pachytene Stage / Q000528:radiation effects*; D013087:Spermatids / Q000378:metabolism* / Q000473:pathology; D013090:Spermatocytes / Q000378:metabolism* / Q000473:pathology
chemical_list: D006570:Heterochromatin; D006657:Histones; C475961:gamma-H2AX protein, mouse