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Malignant transformation of mouse BALB/3T3 cells by polyoma middle T antigen requires epidermal growth factor
Abstract:
The mouse cell line MO-5, which is defective in receptor-binding activity of epidermal growth factor (EGF), is very poorly transformed by polyoma middle T antigen or v-src gene, but activated c-H-ras and v-mos gene can induce the transformation (M. Ono, M. Yakushinji, K. Segawa, and M. Kuwano, Mol. Cell. Biol., 8: 4190-4196, 1988). We established clones of MO-5 expressing a functional EGF receptor (EGF-R) after introduction of the human EGF-R complementary DNA into MO-5 (MNER23 and MNER31), and we also established a clone (BNER4) expressing human EGF-R from the parental cell line, BALB/3T3. MNER23, MNER31, and BNER4 expressed EGF-R activity at about 2- to 6-fold higher levels than did control BALB/3T3 cells. A marked increase in DNA synthesis in response to EGF was observed in these BNER4, MNER23, and MNER31 cell lines compared to BALB/3T3 cells; however, there was little if any increase in DNA synthesis of MO-5 in the presence of EGF. Introduction of the polyoma middle T antigen gene into BALB/3T3, BNER4, MNER23, and MNER31 resulted in the appearance of transformation foci, but MO-5 again showed little response. We purified clones B4-mT-2, M23-mT-1, M23-mT-2, M23-mT-3, and M31-mT-13 from transformation foci of BNER4, MNER23, and MNER31 cells, which were respectively transfected with the middle T antigen. All of the middle T antigen-positive transfectants demonstrated abilities to form both colonies in soft agar and tumors in nude mice. The presence of EGF-R appears to be indispensable for malignant transformation by polyoma middle T antigen.
Insights
The presence of functional epidermal growth factor receptors (EGF-R) is essential for malignant transformation induced by polyoma middle T antigen. Restoring EGF-R in defective cells enabled transformation, highlighting EGF-R
Area of Science:
- Cell Biology
- Molecular Oncology
- Signal Transduction
Background:
- The mouse cell line MO-5 exhibits poor transformation by polyoma middle T antigen due to defective epidermal growth factor (EGF) receptor binding.
- Activated c-H-ras and v-mos genes can induce transformation in MO-5 cells, indicating specific pathway dependencies.
Purpose of the Study:
- To investigate the indispensable role of functional EGF receptors in polyoma middle T antigen-mediated malignant transformation.
- To establish and characterize cell lines with restored or enhanced EGF receptor activity.
Main Methods:
- Genetic engineering of MO-5 cells to express human EGF receptors (creating MNER23 and MNER31 clones).
- Transfection of BALB/3T3 and engineered MO-5 cells with the polyoma middle T antigen gene.
- Assessment of cell transformation, DNA synthesis in response to EGF, colony formation in soft agar, and tumor formation in nude mice.
Main Results:
- Engineered MO-5 cells (MNER23, MNER31) and BNER4 cells showed significantly increased EGF-R activity and DNA synthesis in response to EGF compared to parental MO-5 cells.
- Introduction of polyoma middle T antigen induced transformation foci in cells expressing functional EGF receptors, but not in MO-5 cells.
- All middle T antigen-positive transfectants with functional EGF receptors formed colonies in soft agar and tumors in nude mice.
Conclusions:
- Functional EGF receptors are indispensable for polyoma middle T antigen-induced malignant transformation.
- Restoration of EGF receptor activity in defective cell lines rescues the transformation phenotype.
- This study elucidates a critical requirement for EGF receptor signaling in oncogenesis mediated by specific viral antigens.