Related Experiment Videos
Human mutant cell lines with altered RNA polymerase II
Journal of Cellular Physiology
|November 1, 1982
Summary
Researchers developed an alpha-amanitin-resistant fibrosarcoma cell line. This resistance stems from a stable, dominant mutation altering RNA polymerase II, decreasing its toxin sensitivity.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Alpha-amanitin is a potent toxin that inhibits RNA polymerase II.
- Drug resistance in cancer cells is a significant challenge in chemotherapy.
- Understanding resistance mechanisms is crucial for developing effective treatments.
Purpose of the Study:
- To isolate and characterize a human fibrosarcoma cell line resistant to alpha-amanitin.
- To determine the biochemical basis of this resistance.
Main Methods:
- Ethylmethanesulfonate mutagenesis and stepwise selection were used to generate resistant cells.
- RNA polymerase II was purified from both wild-type and mutant cells.
- The binding affinity of purified RNA polymerase II to alpha-amanitin was assessed.
Main Results:
- A stable, dominant mutation conferring resistance to alpha-amanitin was identified in the HT-1080-6TG-9AM cell line.
- Purified RNA polymerase II from mutant cells exhibited significantly reduced sensitivity (50- to 100-fold) to alpha-amanitin.
- Altered affinity of RNA polymerase II for alpha-amanitin was observed.
Conclusions:
- The biochemical basis for alpha-amanitin resistance in this cell line is an alteration in RNA polymerase II.
- This study provides a model for investigating drug resistance mechanisms involving RNA polymerase II.