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Published on: December 27, 2016
Selective killing induced by an inhibitor of N-linked glycosylation
O Larsson1, M Carlberg, A Zetterberg
1Department of Tumor Pathology, Karolinska Hospital, Stockholm, Sweden.
Abstract:
Treatment with a low dose (0.5 microgram/ml) of tunicamycin (an inhibitor of N-linked glycosylation) blocked the cell cycle progression of both normal Balb/c 3T3 cells (A31) and their SV40-transformed derivatives (SVA31) specifically in early G1 (0-3 h after mitosis). Upon release after an 8-h treatment the A31 cells returned to the cell cycle via a 9-h recovery phase, indicating that they were arrested in G0. The A31 cells were fully viable after this treatment. In contrast, the postmitotic SVA31 cells, which were unable to arrest in G0, did not divide after the removal of tunicamycin. Instead, these cells died but this did not occur until 22-34 h after release from the treatment. SVA31 cells that had passed the postmitotic phase of G1 survived during the parental generation and divided normally. However, a large portion of these cells died during the next cycle, and in total during a 48-h period approximately 50% of the cells were killed as a consequence of an 8-h exposure to tunicamycin. In contrast, treatment with inhibitors of protein synthesis and HMG CoA reductase activity as well as inhibitors of modification of N-linked oligosaccharide chains did not result in cell death.
Insights
Tunicamycin treatment halts cell cycle progression in normal and SV40-transformed cells. Transformed cells exhibit delayed death after tunicamycin removal, unlike normal cells which recover.
Area of Science:
- Cell Biology
- Biochemistry
Background:
- N-linked glycosylation is crucial for protein function and cell cycle regulation.
- SV40 transformation alters cellular responses to cell cycle inhibitors.
Purpose of the Study:
- To investigate the effects of tunicamycin, an N-linked glycosylation inhibitor, on cell cycle progression and viability in normal and SV40-transformed mouse fibroblasts.
- To determine if transformed cells exhibit differential sensitivity to tunicamycin-induced cell cycle arrest compared to normal cells.
Main Methods:
- Balb/c 3T3 cells (A31) and their SV40-transformed counterparts (SVA31) were treated with tunicamycin (0.5 microgram/ml) for 8 hours.
- Cell cycle progression was monitored by flow cytometry after tunicamycin removal.
- Cell viability was assessed over a 48-hour period post-treatment.
Main Results:
- Tunicamycin blocked both normal (A31) and transformed (SVA31) cells in early G1 phase.
- A31 cells recovered and re-entered the cell cycle after treatment, indicating G0 arrest and viability.
- SVA31 cells, unable to arrest in G0, exhibited delayed death (22-34 hours post-release), with ~50% cell death within 48 hours.
- Cells surviving the initial tunicamycin exposure in SVA31 showed increased mortality in subsequent cell cycles.
Conclusions:
- Tunicamycin-induced N-linked glycosylation inhibition causes cell cycle arrest in early G1 for both normal and SV40-transformed fibroblasts.
- SV40 transformation confers a lethal sensitivity to tunicamycin, characterized by delayed cell death and impaired G0 arrest.
- The findings highlight the critical role of N-linked glycosylation in maintaining the viability of transformed cells and suggest potential therapeutic vulnerabilities.
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