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Effects of signalling transduction modulators on the transformed phenotypes in v-H-ras-transformed NIH 3T3 cells
1Institute of Toxicology, College of Medicine, National Taiwan University, Taipei, Republic of China.
Abstract:
Several signalling transduction modulators were used to examine their effects on the morphological changes, foci formation in soft agar and cellular growth in v-H-ras-transformed NIH 3T3 cells. The results from this study showed that specific tyrosine kinase inhibitors (genistein and tyrphostin 23) and cyclic AMP-elevating agents (forskolin and 3-isobutyl-1-methyl-xanthine) could effectively induce differential flat phenotype of v-H-ras transformant at micromolar concentrations. At the same dose range, both signalling modulators also caused a significant suppression of anchorage-independent and cellular growth in the same transformant. By contrast, compound inhibitors such as protein kinase C (staurosporin and H-7), phospholipase A2 (aristolochic acid), phospholipase C (neomycin sulfate) and cyclooxygenase (indomethacin) all did not alter the cellular morphology or foci formation in soft agar, although PKC inhibitors exhibited a slight inhibition on the cellular growth. Based on these observations, we propose that the alterations of protein kinase A or tyrosine kinase-associated signal pathways is necessary and the original cause of the transformation event, but that increase of the activities of protein kinase C, phospholipase C, phospholipase A2 or cyclooxygenase probably is an indirect result of the v-H-ras-mediated transformation.
Insights
Specific tyrosine kinase inhibitors and cyclic AMP-elevating agents reversed v-H-ras transformation. These signaling modulators suppressed cellular growth and induced a flat phenotype, suggesting key roles for protein kinase A and tyrosine kinase pathways.
Area of Science:
- Cell Biology
- Molecular Biology
- Signal Transduction
Background:
- Ras-transformed cells exhibit altered morphology, uncontrolled growth, and anchorage independence.
- Signal transduction pathways play critical roles in cellular transformation and oncogenesis.
Purpose of the Study:
- To investigate the effects of various signaling transduction modulators on v-H-ras-transformed NIH 3T3 cells.
- To identify key signaling pathways involved in the maintenance of the transformed phenotype.
Main Methods:
- Treatment of v-H-ras-transformed NIH 3T3 cells with specific inhibitors and agents.
- Assessment of morphological changes, soft agar colony formation, and cellular growth.
- Evaluation of inhibitors targeting tyrosine kinase, protein kinase A, protein kinase C, phospholipase A2, phospholipase C, and cyclooxygenase.
Main Results:
- Tyrosine kinase inhibitors (genistein, tyrphostin 23) and cyclic AMP-elevating agents (forskolin, 3-isobutyl-1-methyl-xanthine) induced a flat phenotype and suppressed anchorage-independent and cellular growth.
- Inhibitors of protein kinase C, phospholipase A2, phospholipase C, and cyclooxygenase did not significantly alter morphology or foci formation, though PKC inhibitors showed slight growth inhibition.
Conclusions:
- Alterations in protein kinase A or tyrosine kinase-associated pathways are necessary for v-H-ras-mediated transformation.
- Increased activities of protein kinase C, phospholipase C, phospholipase A2, or cyclooxygenase are likely indirect consequences of v-H-ras transformation.