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Establishment of an apoptosis-resistant and growth-controllable cell line by transfecting with inducible antisense
1Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Bunkyo-ku, Tokyo 113, Japan. sarah@bio.t.u-tokyo.ac.jp
Abstract:
F-MEL cells were transfected with the c-jun antisense gene located downstream of a glucocorticoid-inducible MMTV promoter, and the obtained cells were named c-jun AS cells. When the c-jun AS cells were treated with dexamethasone (DEX) in DMEM supplemented with 10% serum, the growth of the cells was completely suppressed for a duration of 16 days with a high cell viability exceeding 86%. The c-jun expression in the c-jun AS cells was suppressed moderately in the absence of DEX and strongly in the presence of DEX. The c-jun AS cells grew well and reached a density of 10(6) cells/mL without supplementation of any serum components. Viability was greater than 80% after the cells had been cultured for 8 days in the absence of DEX. The c-jun AS cells stayed at a constant cell density and high viability above 80% for 8 days when they were cultured in the presence of DEX under serum deprivation. In contrast, the wild type F-MEL cells were unable to grow and died by apoptosis in 3 days under serum deprivation. Internucleosomal cleavage of DNA, a landmark of apoptosis, was clearly detectable. Thus the c-jun AS cell line that is resistant to apoptosis induced by serum deprivation and can reversibly and viably be growth-arrested was established. A dual-signal model was proposed to explain the experimental result, the interlinked regulation of apoptosis, and growth by c-jun.
Insights
A novel cell line, c-jun AS cells, demonstrates resistance to apoptosis and reversible growth arrest. This discovery offers new insights into the regulation of cell growth and apoptosis by c-jun.
Area of Science:
- Molecular Biology
- Cell Biology
- Cancer Research
Background:
- The c-jun gene plays a critical role in regulating cell proliferation and apoptosis.
- Understanding c-jun's function is crucial for developing targeted cancer therapies.
- Developing cell models to study gene function is essential in molecular biology.
Purpose of the Study:
- To establish a cell line with inducible suppression of c-jun expression.
- To investigate the role of c-jun in apoptosis and cell growth regulation.
- To create a tool for studying reversible growth arrest and apoptosis resistance.
Main Methods:
- Transfection of F-MEL cells with a c-jun antisense gene under a glucocorticoid-inducible promoter (MMTV).
- Treatment with dexamethasone (DEX) to induce c-jun suppression.
- Cell culture under various conditions (serum-supplemented, serum-deprived, presence/absence of DEX) to assess growth and viability.
- Analysis of DNA fragmentation to detect apoptosis.
Main Results:
- Established c-jun AS cells exhibited suppressed c-jun expression upon DEX treatment.
- c-jun AS cells showed complete growth suppression and high viability (>86%) for 16 days with DEX.
- c-jun AS cells demonstrated resistance to apoptosis induced by serum deprivation, maintaining high viability for 8 days.
- Wild-type F-MEL cells underwent apoptosis and died within 3 days under serum deprivation.
Conclusions:
- The established c-jun AS cell line is resistant to serum deprivation-induced apoptosis.
- This cell line allows for reversible and viable growth arrest, controlled by DEX.
- A dual-signal model suggests c-jun interlinked regulation of apoptosis and growth.