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Activation of a histone H1 kinase by tyrosine phosphorylation in v-src-transformed fibroblasts
D W Sternberg1, G Scholz, Y Fukui
1Laboratory of Molecular Oncology, Rockefeller University, New York, NY 10021.
Abstract:
Using anti-phosphotyrosine immunoaffinity chromatography, we have searched for serine/threonine kinases that are directly regulated by tyrosine phosphorylation in v-src-transformed rat 3Y1 fibroblasts. Tyrosine phosphoprotein preparations from v-src-transformed cells contain a kinase activity that phosphorylates histone H1 in vitro on serine residues and this activity is present at a 20-fold greater level than that in parental cell preparations. This activity elutes from a MonoQ FPLC column as a single peak and gel filtration chromatography suggests that the kinase has a molecular mass of approximately 55 kDa. Tyrosine phosphatase treatment inactivates the histone H1 kinase and this result indicates that the specific activity of the kinase is regulated by tyrosine phosphorylation. Experiments with cells transformed with a temperature-sensitive mutant of the v-src oncogene demonstrate that the tyrosine phosphorylation of the histone H1 kinase is an early event in v-src transformation. The kinase is distinct from known cdc2 family members that contain the PSTAIR motif, because the kinase can be separated almost completely from these proteins by immunoprecipitation with an antibody against p34cdc2. The profile of antibody reactivity and sensitivity to modulators of protein kinases suggests that this activity is distinct from known second messenger-regulated kinases and from previously characterized MAP kinases.
Insights
Researchers identified a novel histone H1 kinase regulated by tyrosine phosphorylation in v-src-transformed cells. This kinase activity is significantly elevated and crucial for early v-src transformation events.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- v-src oncogene is known to induce cellular transformation.
- Tyrosine phosphorylation plays a critical role in regulating protein function during cell signaling.
Purpose of the Study:
- To identify serine/threonine kinases directly regulated by tyrosine phosphorylation in v-src-transformed cells.
- To characterize the properties and role of such a kinase in v-src-mediated transformation.
Main Methods:
- Anti-phosphotyrosine immunoaffinity chromatography was used to isolate tyrosine-phosphorylated proteins.
- Histone H1 kinase activity was assayed in vitro.
- MonoQ FPLC and gel filtration chromatography were employed for protein purification and size estimation.
- Experiments with temperature-sensitive v-src mutants and tyrosine phosphatase treatment were performed.
Main Results:
- A novel histone H1 kinase activity was identified in v-src-transformed cells, showing a 20-fold increase compared to parental cells.
- The kinase has an estimated molecular mass of 55 kDa and its activity is dependent on tyrosine phosphorylation.
- This tyrosine phosphorylation is an early event in v-src transformation.
- The kinase is distinct from known cdc2 family members and other regulated kinases like MAP kinases.
Conclusions:
- A novel serine/threonine kinase regulated by tyrosine phosphorylation is involved in v-src-induced cell transformation.
- This kinase represents a potential new target for understanding and potentially treating cancers driven by v-src or related tyrosine kinases.