Related Experiment Videos
Induction of murine teratocarcinoma cell differentiation by suppression of poly(ADP-ribose) synthesis
Abstract:
Poly(ADP-ribose) synthesizing activity in mouse teratocarcinoma EC-A1 cells decreased markedly during differentiation induced by retinoic acid; the activities assayed in permeabilized cells decreased to 25% and 10% of the activity of control (uninduced cells) 2 and 3 days, respectively, after the addition of 0.1 microM retinoic acid to the culture medium. This change preceded changes in morphology and DNA synthesis, which became prominent after 4 days. The decrease in poly(ADP-ribose) synthesizing activity appeared to be caused by a diminution of the synthetase protein and not by a decrease in its catalytic activity, because the full activity disclosed by DNase I treatment decreased in parallel, albeit at about 20 times higher levels. When 8 mM 3-aminobenzamide or 10 mM nicotinamide, specific inhibitors of poly(ADP-ribose) synthetase, was added to the culture medium, the cells underwent differentiation after 7-9 days. An analogue, 3-aminobenzoic acid, which is not inhibitory to the synthetase, induced differentiation much less efficiently than did 3-aminobenzamide, and the effect of 3-aminobenzoic acid appeared to be ascribable to its potent cytotoxicity. Immunohistochemical analysis using anti-poly(ADP-ribose) antibody confirmed the marked reduction in poly(ADP-ribose) synthesizing activity in nuclei of the cells treated with retinoic acid or 3-aminobenzamide but not with 3-aminobenzoic acid. These results suggest that a decrease in poly(ADP-ribose) synthesis triggers differentiation of teratocarcinoma cells.
Insights
Differentiation of mouse teratocarcinoma cells is triggered by a decrease in poly(ADP-ribose) synthesis. This reduction in poly(ADP-ribose) synthesizing activity precedes morphological changes and DNA synthesis during retinoic acid-induced differentiation.
Area of Science:
- Cell Biology
- Biochemistry
- Developmental Biology
Background:
- Poly(ADP-ribose) synthesis is a crucial cellular process.
- Teratocarcinoma cells provide a model for studying differentiation.
- The role of poly(ADP-ribose) in cell differentiation is not fully understood.
Purpose of the Study:
- To investigate the role of poly(ADP-ribose) synthesis in retinoic acid-induced differentiation of mouse teratocarcinoma cells.
- To determine whether a decrease in poly(ADP-ribose) synthesis triggers differentiation.
Main Methods:
- Assaying poly(ADP-ribose) synthesizing activity in permeabilized cells.
- Monitoring cell morphology and DNA synthesis.
- Using inhibitors of poly(ADP-ribose) synthetase (3-aminobenzamide, nicotinamide) and a non-inhibitory analogue (3-aminobenzoic acid).
- Immunohistochemical analysis using anti-poly(ADP-ribose) antibody.
Main Results:
- Retinoic acid treatment markedly decreased poly(ADP-ribose) synthesizing activity in EC-A1 cells, preceding morphological and DNA synthesis changes.
- The decrease in activity was attributed to reduced synthetase protein, not altered catalytic efficiency.
- Inhibitors of poly(ADP-ribose) synthetase induced differentiation, while a non-inhibitory analogue showed less effect.
- Immunohistochemistry confirmed reduced poly(ADP-ribose) levels in differentiating cells.
Conclusions:
- A decrease in poly(ADP-ribose) synthesis appears to be a trigger for teratocarcinoma cell differentiation.
- Poly(ADP-ribose) synthetase activity is modulated during retinoic acid-induced differentiation.
- Targeting poly(ADP-ribose) synthesis may offer a novel approach to controlling cell differentiation.