Replication factories and nuclear bodies: the ultrastructural characterization of replication sites during the cell cycle.

Journal article

Sites of replication in synchronized HeLa cells were visualized by light and electron microscopy; cells were permeabilized and incubated with biotin-16-dUTP, and incorporation sites were immunolabelled. Electron microscopy of thick resinless sections from which approximately 90% chromatin had been removed showed that most DNA synthesis occurs in specific dense structures (replication factories) attached to a diffuse nucleoskeleton. These factories appear at the end of G1-phase and quickly become active; as S-phase progresses, they increase in size and decrease in number like sites of incorporation seen by light microscopy. Electron microscopy of conventional thin sections proved that these factories are a subset of nuclear bodies; they changed in the same characteristic way and contained DNA polymerase alpha and proliferating cell nuclear antigen. As replication factories can be observed and labelled in non-permeabilized cells, they cannot be aggregation artifacts. Some replication occurs outside factories at discrete sites on the diffuse skeleton; it becomes significant by mid S-phase and later becomes concentrated beneath the lamina.

Authors: Hozák, P; Jackson, D; Cook, P

• Publication status: Published
• Journal: Journal of cell science
• Volume: 107 ( Pt 8)
• Pages: 2191-2202
• Publication date: 1994-08-01
• EISSN: 1477-9137
• ISSN: 0021-9533
• Language: English
• Keywords: Biotin; Proliferating Cell Nuclear Antigen; Cell Nucleus; Deoxyuracil Nucleotides; Nuclear Matrix; DNA Replication; Cell Membrane Permeability; Chromatin; Models, Genetic; DNA Polymerase II; Chromosomes; Microscopy, Immunoelectron; Hela Cells; S Phase; Humans