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Tob, a novel protein that interacts with p185erbB2, is associated with anti-proliferative activity
S Matsuda1, J Kawamura-Tsuzuku, M Ohsugi
1Department of Oncology, University of Tokyo, Japan.
Abstract:
We have molecularly cloned a cDNA for a novel protein termed Tob (Transducer of ErbB-2) that interacts with the c-erbB-2 gene product p185erbB2. Nucleotide sequencing reveals that the Tob protein is a 45 kDa protein that does not contain either SH2 (Src Homology 2) or SH3 domain but is homologous to the previously characterized anti-proliferative gene product BTG-1 at its amino-terminal half. The carboxyl-terminal half of Tob is characterized by the presence of a sequence rich in proline and glutamine and shows no homology to known proteins. Like BTG-1, exogenously expressed Tob is able to suppress growth of NIH3T3 cells, but the growth suppression is hampered by the presence of kinase-active p185erbB2. By using the GST-Tob protein that contains either full length or amino-terminal half of Tob, we show that the carboxyl-terminal half of Tob is relevant to its interaction with p185erbB2. Furthermore, we could co-immunoprecipitate the Tob protein with anti-ErbB-2 antibody, and reciprocally the p185erbB2 with anti-Tob antibodies. These data suggest that p185erbB2 negatively regulates the Tob-mediated anti-proliferative pathway through its interaction with Tob, resulting possibly in growth stimulation by p185erbB2. Finally, expression of the Tob mRNA is observed in various cell types and is not correlated with expression of c-erbB-2, suggesting that other receptor-type protein-tyrosine kinases are also involved in the Tob-mediated regulation of cell growth.
Insights
We identified Tob, a novel protein that interacts with c-erbB-2. Tob suppresses cell growth, but this effect is reduced by active c-erbB-2, suggesting c-erbB-2 regulates Tob's anti-proliferative pathway.
Area of Science:
- Molecular biology
- Cell signaling
- Oncology
Background:
- The c-erbB-2 proto-oncogene product p185erbB2 is a receptor-type tyrosine kinase implicated in cell growth and cancer.
- Understanding proteins that interact with p185erbB2 is crucial for elucidating cell growth regulation.
- The anti-proliferative gene BTG-1 shares homology with novel proteins.
Purpose of the Study:
- To identify and characterize novel proteins interacting with the c-erbB-2 gene product p185erbB2.
- To investigate the functional role of the novel protein Tob (Transducer of ErbB-2) in cell growth regulation.
- To determine the interaction domains and regulatory mechanisms between Tob and p185erbB2.
Main Methods:
- Molecular cloning and nucleotide sequencing of Tob cDNA.
- Expression of recombinant Tob protein and analysis of its effect on NIH3T3 cell proliferation.
- Co-immunoprecipitation assays to confirm the interaction between Tob and p185erbB2.
- Analysis of Tob mRNA expression in various cell types.
Main Results:
- A novel protein, Tob (Transducer of ErbB-2), was cloned and characterized. Tob is 45 kDa, homologous to BTG-1 in its N-terminal half, and interacts with p185erbB2 via its C-terminal half.
- Exogenous Tob expression suppressed NIH3T3 cell growth, an effect diminished by kinase-active p185erbB2.
- Co-immunoprecipitation confirmed the physical interaction between Tob and p185erbB2.
- Tob mRNA expression was detected in various cell types and was not correlated with c-erbB-2 expression.
Conclusions:
- p185erbB2 negatively regulates the Tob-mediated anti-proliferative pathway through direct interaction, potentially leading to growth stimulation.
- The carboxyl-terminal half of Tob is essential for its interaction with p185erbB2.
- Tob's regulation of cell growth may involve other receptor-type protein-tyrosine kinases besides c-erbB-2.