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Differential cell cycle modulation of human DNA glycosylases against oxidized pyrimidines
1Department of Pathology, Temple University School of Medicine, Philadelphia, PA 19140.
Abstract:
Cellular DNA is continuously subject to damages by both endogenous and exogenous oxidizing agents. Excision repair of oxidized bases in human cells is initiated by DNA glycosylases which remove them from DNA. 5-Hydroxymethyluracil-DNA glycosylase excises 5-hydroxymethyluracil from DNA. A different enzyme, termed a redoxyendonuclease, has glycosylase activity against many modified DNA pyrimidines. The regulation of these enzymes in proliferating human cells was examined. Both glycosylases were assayed in serum-stimulated WI-38 cells by measurements of direct release of modified free bases from their respective DNA substrates. There was no significant variation of 5-hydroxymethyluracil-DNA glycosylase activity during the cell cycle. However, the glycosylic activity of the redoxyendonuclease was stimulated with DNA synthesis. This activity again increased at the beginning of a second cell cycle. Therefore, the glycosylases that initiate excision repair of oxidized DNA are subject to different controls during the cell cycle.
Insights
DNA repair enzymes, 5-hydroxymethyluracil-DNA glycosylase and redoxyendonuclease, show distinct cell cycle regulation in human cells. Redoxyendonuclease activity increases with DNA synthesis, unlike the other enzyme.
Area of Science:
- Molecular Biology
- Biochemistry
- Cell Biology
Background:
- Cellular DNA is constantly damaged by oxidizing agents, necessitating repair mechanisms.
- DNA glycosylases initiate excision repair by removing oxidized bases from DNA.
- Key enzymes include 5-hydroxymethyluracil-DNA glycosylase and a redoxyendonuclease with broad pyrimidine damage activity.
Purpose of the Study:
- To investigate the cell cycle regulation of DNA glycosylases involved in repairing oxidized DNA bases in human cells.
- To compare the regulation of 5-hydroxymethyluracil-DNA glycosylase and redoxyendonuclease activity during proliferation.
Main Methods:
- Assaying glycosylase activity in serum-stimulated WI-38 human cells.
- Measuring the direct release of modified free bases from DNA substrates.
- Analyzing enzyme activity across different phases of the cell cycle.
Main Results:
- 5-Hydroxymethyluracil-DNA glycosylase activity showed no significant variation throughout the cell cycle.
- Redoxyendonuclease activity was significantly stimulated during DNA synthesis.
- This redoxyendonuclease activity further increased at the onset of a second cell cycle.
Conclusions:
- DNA repair glycosylases initiating oxidized base excision exhibit differential regulation during the cell cycle.
- Redoxyendonuclease activity is linked to DNA replication, suggesting a role in repair during S-phase.
- These findings highlight distinct control mechanisms for DNA repair enzymes in proliferating cells.