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Expression cDNA cloning of a transforming gene encoding the wild-type G alpha 12 gene product
A M Chan1, T P Fleming, E S McGovern
1Laboratory of Cellular and Molecular Biology, National Cancer Institute, Bethesda, Maryland 20892.
Abstract:
Using an expression cDNA cloning approach, we examined human tumor cell lines for novel oncogenes that might evade detection by conventional techniques. We isolated a transforming sequence that was highly efficient in transforming NIH 3T3 mouse fibroblasts. DNA sequence analysis identified the gene as the human homolog of a recently cloned alpha subunit of mouse GTP-binding protein G alpha 12. NIH 3T3 cells transfected with G alpha 12 cDNA grew in soft agar and were tumorigenic in nude mice. There were no apparent mutations in the cloned cDNA in comparison with a G alpha 12 cDNA clone isolated from a normal human epithelial cell library, implying that overexpression alone was sufficient to cause NIH 3T3 cell transformation. The observed altered growth properties mediated by G alpha 12 showed a certain degree of dependency on serum factors, and its mitogenic potential was also potently inhibited by suramin treatment.
Insights
Researchers identified a novel oncogene, G alpha 12, in human tumors. Overexpression of this GTP-binding protein subunit transforms mouse cells, indicating its role in cancer development.
Area of Science:
- Molecular Biology
- Oncology
- Cell Biology
Background:
- Identifying novel oncogenes is crucial for understanding cancer.
- Conventional techniques may miss certain transforming genes.
- GTP-binding proteins play roles in cell signaling and growth.
Purpose of the Study:
- To isolate and characterize novel oncogenes from human tumor cell lines.
- To determine if the human homolog of mouse G alpha 12 has oncogenic properties.
- To investigate the mechanism of transformation mediated by G alpha 12.
Main Methods:
- Expression cDNA cloning from human tumor cell lines.
- NIH 3T3 mouse fibroblast transformation assays.
- DNA sequence analysis.
- Soft agar colony formation assays.
- Tumorigenicity studies in nude mice.
Main Results:
- A transforming sequence was isolated and identified as human G alpha 12.
- NIH 3T3 cells transfected with G alpha 12 cDNA exhibited anchorage-independent growth and tumorigenicity.
- No mutations were found in the cloned G alpha 12 cDNA, suggesting overexpression causes transformation.
- G alpha 12-mediated transformation showed serum dependency and was inhibited by suramin.
Conclusions:
- Human G alpha 12 acts as an oncogene, capable of transforming NIH 3T3 cells through overexpression.
- G alpha 12 plays a role in altered cell growth and mitogenesis.
- Further research into G alpha 12's role in human tumorigenesis is warranted.