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Differential expression of tropomyosin forms in the microfilaments isolated from normal and transformed rat cultured
Abstract:
Using a newly developed method for microfilament isolation (Matsumura, F., Yamashiro-Matsumura, S. and Lin, J. J.-C. (1983) J. Biol. Chem. 258, 6636-6644), we have analyzed protein composition of microfilaments in "normal" and transformed rat tissue culture cells. They include REF-52 (an established rat embryo cell line) cells, REF-52 transformed by DNA viruses (SV40 or adenovirus type 5), normal rat kidney cells, and normal rat kidney cells transformed by RNA viruses (Kirsten or Rous sarcoma virus). Microfilaments from normal rat culture cells contain three major tropomyosins (apparent Mr = 40,000, 36,500, and 32,400) and two relatively minor tropomyosins (apparent Mr = 35,000 and 32,000). In transformed cells the levels of one or two of the major tropomyosins (Mr = 40,000 and 36,500) are decreased and the levels of one or both of the minor tropomyosins (Mr = 35,000 and 32,000) are increased. These changes in tropomyosin patterns were also observed in temperature shift experiments with rat-1 cells transformed with a Rous sarcoma virus mutant, temperature-sensitive for transformation. Cell-free translation of whole cell mRNA generated similar tropomyosin patterns on two-dimensional gels, suggesting that changes in the pattern of tropomyosin expression were largely effected at the level of RNA rather than by post-translational modification. Such changes in the tropomyosin composition of microfilaments were consistently found to accompany the various morphological alterations associated with transformation. We suggest that alterations in the pattern of tropomyosin expression are involved in, or cause, rearrangement of stress fibers and that this may be responsible (in part) for morphological transformation.
Insights
Cancer cell transformation involves changes in microfilament protein composition. Key tropomyosin proteins decrease while minor ones increase, suggesting RNA-level regulation impacts cell structure and morphology.
Area of Science:
- Cell Biology
- Biochemistry
- Molecular Biology
Background:
- Microfilaments are crucial for cell structure and motility.
- Transformation of cells by viruses alters their morphology and behavior.
- Tropomyosin is a key protein component of microfilaments.
Purpose of the Study:
- To analyze the protein composition of microfilaments in normal and virally transformed rat cells.
- To investigate changes in tropomyosin expression during cellular transformation.
- To determine if these changes are regulated at the RNA level.
Main Methods:
- Developed a new method for microfilament isolation.
- Analyzed microfilament protein composition using gel electrophoresis.
- Performed cell-free translation of mRNA to assess RNA-level regulation.
- Utilized temperature-shift experiments with temperature-sensitive viral mutants.
Main Results:
- Normal rat cells contain specific major and minor tropomyosin isoforms.
- Transformed cells exhibit decreased levels of major tropomyomysins and increased levels of minor tropomyosins.
- These tropomyosin pattern changes were observed across various viral transformation models (DNA and RNA viruses).
- Changes in tropomyosin expression were largely regulated at the RNA level, not by post-translational modification.
- Tropomyosin alterations correlated with morphological changes associated with transformation.
Conclusions:
- Alterations in tropomyosin composition of microfilaments accompany cellular transformation.
- Changes in tropomyosin expression patterns are implicated in the rearrangement of stress fibers.
- These microfilament changes likely contribute to the morphological transformation of cells.