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Activation of Ty transposition by mutagens
1Department of Cell Biology, Room 698, School of Medicine, 550 First Avenue, New York University, New York, NY 10016, USA. stale101@popmail.med.nyu.edu
Abstract:
The induction of Ty1 transposition by mutagens (MMS and 4NQO) in asynchronous cultures and cells blocked in G1 and G2/M suggested G1 dependence of activation of Ty1 element by DNA damage. Northern blot analysis revealed immediate five-fold increase in levels of Ty1 transcript after 20min incubation of cells with 1 microg/ml 4NQO and four-fold increase in Ty1 RNA after treatment the cells with 0.1% MMS. Western blot analysis showed no difference in TyA protein in treated and untreated with mutagen cells. Quantitative mutagenicity assay and Northern blot analysis demonstrated dependence of induction of Ty1 element by DNA-damaging agents on the function of RAD9 gene and independence on DUN1 gene.
Insights
DNA damage activates Ty1 transposition in G1 phase, requiring RAD9 but not DUN1. This DNA damage response increases Ty1 RNA levels but not TyA protein, indicating post-transcriptional regulation.
Area of Science:
- Molecular Biology
- Genetics
- Cell Cycle Regulation
Background:
- Ty1 elements are retrotransposons in yeast that can influence genome stability.
- DNA damage can trigger various cellular responses, including alterations in gene expression and transposition.
- The cell cycle plays a crucial role in regulating DNA replication, repair, and genome integrity.
Purpose of the Study:
- To investigate the role of cell cycle phase in the induction of Ty1 transposition by DNA-damaging agents.
- To elucidate the molecular mechanisms underlying Ty1 element activation in response to genotoxic stress.
- To identify key genes involved in the DNA damage-induced regulation of Ty1 transposition.
Main Methods:
- Treatment of yeast cultures with DNA mutagens (MMS and 4NQO) at different cell cycle phases (asynchronous, G1, G2/M).
- Northern blot analysis to quantify Ty1 RNA transcript levels.
- Western blot analysis to assess TyA protein levels.
- Quantitative mutagenicity assays and Northern blot analysis to evaluate gene function dependence (RAD9 and DUN1).
Main Results:
- Induction of Ty1 transposition by mutagens is dependent on the G1 phase of the cell cycle.
- Treatment with 4NQO and MMS significantly increased Ty1 RNA levels (five-fold and four-fold, respectively) within 20 minutes.
- No significant changes in TyA protein levels were observed between mutagen-treated and untreated cells.
- Induction of Ty1 transposition by DNA-damaging agents requires the RAD9 gene but is independent of the DUN1 gene.
Conclusions:
- DNA damage-induced activation of Ty1 transposition is a G1-dependent process.
- The regulation of Ty1 transposition by DNA damage primarily occurs at the transcriptional level.
- The RAD9 checkpoint pathway is essential for mediating the induction of Ty1 transposition in response to DNA damage.