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Focus Formation: A Cell-based Assay to Determine the Oncogenic Potential of a Gene
Published on: December 31, 2014
Increased cell-substrate adhesion accompanies conditional reversion to the normal phenotype in
D K Shumaker1, M D Sklar, E V Prochownik
1Department of Pathology, University of Michigan Medical School, Ann Arbor 48109.
Experimental Cell Research
|October 1, 1994
Summary
Depleting c-myc protein in ras-transformed cells reversed malignancy and increased cell adhesion. This change was linked to more surface fibronectin, suggesting a role in regaining a nonmalignant phenotype.
Area of Science:
- Oncology
- Cell Biology
- Molecular Biology
Background:
- Ras-transformed NIH-3T3 cells exhibit malignant characteristics.
- Depletion of c-myc protein previously reversed these malignant traits.
- The role of c-myc in cell adhesion was not fully understood.
Purpose of the Study:
- To investigate the effect of c-myc depletion on the in vitro adhesive behavior of ras-transformed NIH-3T3 cells.
- To examine fibronectin biosynthesis and cell surface fibronectin levels in c-myc depleted cells.
Main Methods:
- Utilized antisense transfection to deplete c-myc protein in ras-transformed NIH-3T3 cells.
- Assessed cell attachment to fibronectin-coated dishes.
- Measured resistance to trypsin/EDTA-mediated cell release from the substratum.
- Quantified fibronectin biosynthesis and analyzed cell surface fibronectin via immunofluorescence staining.
Main Results:
- Cells with depleted c-myc showed no change in initial attachment to fibronectin.
- These cells exhibited significantly increased resistance to detachment from the substratum.
- No overall change in fibronectin biosynthesis was observed.
- Immunofluorescence revealed increased surface fibronectin in c-myc depleted cells.
Conclusions:
- Conditional depletion of c-myc protein in ras-transformed cells leads to increased cell-substrate adhesion.
- This enhanced adhesion is associated with increased surface fibronectin.
- These findings suggest c-myc plays a role in regulating cell adhesion and the nonmalignant phenotype.
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