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Biochemical and functional characterization of the murine ros protooncogene
D Riethmacher1, O Langholz, S Gödecke
1Max-Delbrück-Laboratorium in der Max-Planck-Gesellschaft, Köln, Germany.
Abstract:
The ros gene was originally found because it can, when mutated, induce malignant transformation. The protooncogene encodes an orphan receptor tyrosine kinase. We report here the isolation and characterization of the mouse c-ros cDNA and, in addition, the biochemical characterization of the receptor. Both, the endogenous c-ros protein from embryonal tissues and the recombinant protein are glycosylated molecules with an apparent molecular weight of 260,000. Pulse-chase analysis in Sf9 cells demonstrates that the c-ros protein is synthesized as a single chain, uncleaved molecule. Since the specific ligand of c-ros is not known, a hybrid receptor (trk/c-ros) which transmits c-ros-specific signals in response to nerve growth factor (NGF) was used to study the biological activities. In NIH3T3 cells, this trk/c-ros hybrid induces growth, a fusiform cell shape, and loss of contact inhibition of growth. However, the active hybrid receptor cannot replace IL-3 as survival factor in 32D myeloid cells. Compared to other receptors, the active c-ros tyrosine kinase domain displays thus overlapping, but not identical signalling specificities.
Insights
The ros protooncogene encodes a receptor tyrosine kinase. Researchers characterized the mouse c-ros protein, revealing its glycosylation and signaling specificities, which overlap but differ from other receptors.
Area of Science:
- Molecular Biology
- Cell Signaling
- Oncology
Background:
- The ros gene, a protooncogene, encodes an orphan receptor tyrosine kinase implicated in malignant transformation.
- Understanding the c-ros protein's function is crucial for its role in cell growth and cancer.
Purpose of the Study:
- To isolate and biochemically characterize the mouse c-ros cDNA and its encoded protein.
- To investigate the biological activities and signaling specificities of the c-ros receptor tyrosine kinase.
Main Methods:
- Isolation and characterization of mouse c-ros cDNA.
- Biochemical analysis of endogenous and recombinant c-ros proteins, including glycosylation and molecular weight determination.
- Pulse-chase analysis in Sf9 cells to study protein synthesis.
- Utilizing a trk/c-ros hybrid receptor to study c-ros signaling in response to nerve growth factor (NGF).
Main Results:
- The c-ros protein is a glycosylated molecule with an apparent molecular weight of 260,000.
- Pulse-chase experiments showed c-ros is synthesized as a single, uncleaved chain.
- The trk/c-ros hybrid induced cell growth, altered morphology, and loss of contact inhibition in NIH3T3 cells.
- The c-ros kinase domain exhibited overlapping but distinct signaling specificities compared to other receptors, and could not substitute for IL-3 in 32D myeloid cells.
Conclusions:
- The mouse c-ros receptor tyrosine kinase is a glycosylated protein with unique signaling properties.
- Its signaling pathways partially overlap with other tyrosine kinases but possess distinct functional outcomes.
- Further research into c-ros signaling may elucidate its role in normal development and oncogenesis.