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Talin is phosphorylated on tyrosine in chicken embryo fibroblasts transformed by Rous sarcoma virus
Abstract:
We have examined the extent of tyrosine phosphorylation of talin, a component of the cytoskeleton localized in the focal adhesions and, therefore, a potential substrate of p60v-src, the transforming protein of Rous sarcoma virus. p60v-src is a tyrosine kinase that induces high levels of phosphotyrosine and the disorganization of the cytoskeleton in transformed cells. With a polyclonal antibody utilized in a previous study [Maher, P. A., Pasquale, E. B., Wang, J. Y. J. & Singer, S. J. (1985) Proc. Natl. Acad. Sci. USA 82, 6576-6580] for the detection of tyrosine-phosphorylated proteins, we have detected phosphotyrosine residues in talin molecules immunoprecipitated from Rous sarcoma virus-transformed, but not normal, chicken embryo fibroblasts. Phospho amino acid analysis of talin from the infected cells confirmed the presence of phosphotyrosine, in addition to phosphoserine and phosphothreonine. The extent of tyrosine modification in talin was compared to that in vinculin, the other focal adhesion component previously found to contain enhanced levels of phosphotyrosine in various retrovirus-transformed cells. A considerably (3 times) larger fraction of the talin than of the vinculin molecules was found to be phosphorylated on tyrosine. The phosphorylation of talin on tyrosine may be crucial for the expression of the abnormal morphology characteristic of cells transformed by Rous sarcoma virus.
Insights
Tyrosine phosphorylation of talin, a cytoskeletal protein, is significantly increased in Rous sarcoma virus-transformed cells. This enhanced tyrosine modification of talin may drive the abnormal cell morphology seen in transformed cells.
Area of Science:
- Cell Biology
- Virology
- Biochemistry
Background:
- p60v-src, a Rous sarcoma virus protein, is a tyrosine kinase.
- p60v-src induces cytoskeletal disorganization and increased phosphotyrosine levels in transformed cells.
- Talin is a cytoskeletal component found in focal adhesions.
Purpose of the Study:
- To investigate the tyrosine phosphorylation of talin.
- To determine if talin is a substrate of p60v-src.
- To compare talin phosphorylation to vinculin phosphorylation in transformed cells.
Main Methods:
- Immunoprecipitation of talin from normal and Rous sarcoma virus-transformed chicken embryo fibroblasts.
- Detection of phosphotyrosine using a specific polyclonal antibody.
- Phospho amino acid analysis of talin.
- Comparison of talin and vinculin tyrosine phosphorylation levels.
Main Results:
- Phosphotyrosine residues were detected in talin immunoprecipitated from transformed cells, but not normal cells.
- Phospho amino acid analysis confirmed phosphotyrosine in talin, alongside phosphoserine and phosphothreonine.
- Talin exhibited approximately three times more tyrosine phosphorylation than vinculin.
Conclusions:
- Talin is tyrosine-phosphorylated in Rous sarcoma virus-transformed cells.
- The extent of talin tyrosine phosphorylation is significantly higher than that of vinculin.
- Talin tyrosine phosphorylation may be essential for the abnormal morphology of Rous sarcoma virus-transformed cells.