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Transformation by ras suppresses expression of the neurotrophic growth factor pleiotrophin
1Division of Cellular and Molecular Biology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA. michael_corbley@dfci.harvard.edu
Abstract:
An 18-kDa protein (p18) was detected in lysates and conditioned medium from contact-arrested NIH 3T3 fibroblasts, but was not detected when the cells were transformed by the oncogene ras. Analysis of transformation-defective cell clones generated after mutagenesis of the ras-retroviral vector used to transduce the ras gene showed an inverse correlation between p18 expression and the degree of transformation. p18 expression was high in non-transformed clones, intermediate in a partially transformed clone, undetectable in fully transformed clones, and detectable only at the non-permissive temperature in a clone which was cold-sensitive for ras transformation. In non-transformed cells, p18 expression varied with the degree of confluence. It was almost undetectable in medium from sparse, proliferating cells, but increased as the cells approached confluence and peaked 2-4 days after confluence. Microsequencing of partially purified p18 identified it as the developmentally regulated neurotrophic factor pleiotrophin. In further experiments, pleiotrophin was undetectable or almost undetectable in medium from fully transformed cells expressing the oncogenes v-src, truncated c-raf, activated c-fms, or polyomavirus middle tumor antigen; it was low but easily detectable in medium from SV40 large tumor antigen-expressing cells, which form soft agar colonies but not foci. Thus, pleiotrophin expression in NIH 3T3 cells is associated with quiescence, and suppression of pleiotrophin is related to oncogenic transformation.
Insights
Pleiotrophin (p18) expression is high in quiescent NIH 3T3 fibroblasts but suppressed by oncogenic transformation. This protein
Area of Science:
- Cell Biology
- Molecular Biology
- Oncology
Background:
- Contact-arrested NIH 3T3 fibroblasts express an 18-kDa protein (p18).
- p18 expression is inversely correlated with the degree of oncogenic transformation.
- p18 was identified as pleiotrophin, a neurotrophic factor.
Purpose of the Study:
- To investigate the relationship between pleiotrophin expression and oncogenic transformation in NIH 3T3 cells.
- To determine how different oncogenes affect pleiotrophin levels.
- To understand the role of pleiotrophin in cell quiescence and transformation.
Main Methods:
- Detection of p18 in cell lysates and conditioned medium.
- Analysis of p18 expression in transformation-defective cell clones.
- Microsequencing to identify p18.
- Assessing pleiotrophin levels in cells expressing various oncogenes (ras, v-src, c-raf, c-fms, polyomavirus middle tumor antigen, SV40 large tumor antigen).
Main Results:
- p18 (pleiotrophin) was detected in contact-arrested, non-transformed NIH 3T3 cells.
- Pleiotrophin expression decreased with increasing oncogenic transformation.
- Pleiotrophin levels were high in confluent, quiescent cells and low in sparse, proliferating cells.
- Expression of ras, v-src, c-raf, c-fms, and polyomavirus middle tumor antigen suppressed pleiotrophin.
- SV40 large tumor antigen partially suppressed pleiotrophin.
Conclusions:
- Pleiotrophin expression is associated with cellular quiescence in NIH 3T3 fibroblasts.
- Suppression of pleiotrophin is linked to oncogenic transformation.
- Pleiotrophin may play a role in maintaining the quiescent state and its suppression contributes to transformation.