The SH2 domain protein GRB-7 is co-amplified, overexpressed and in a tight complex with HER2 in breast cancer
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1New York University Medical Center, Department of Pharmacology, Kaplan Cancer Center, NY 10016.
Abstract:
SH2 domain proteins are important components of the signal transduction pathways activated by growth factor receptor tyrosine kinases. We have been cloning SH2 domain proteins by bacterial expression cloning using the tyrosine phosphorylated C-terminus of the epidermal growth factor receptor as a probe. One of these newly cloned SH2 domain proteins, GRB-7, was mapped on mouse chromosome 11 to a region which also contains the tyrosine kinase receptor, HER2/erbB-2. The analogous chromosomal locus in man is often amplified in human breast cancer leading to overexpression of HER2. We find that GRB-7 is amplified in concert with HER2 in several breast cancer cell lines and that GRB-7 is overexpressed in both cell lines and breast tumors. GRB-7, through its SH2 domain, binds tightly to HER2 such that a large fraction of the tyrosine phosphorylated HER2 in SKBR-3 cells is bound to GRB-7. GRB-7 can also bind tyrosine phosphorylated SHC, albeit at a lower affinity than GRB2 binds SHC. We also find that GRB-7 has a strong similarity over > 300 amino acids to a newly identified gene in Caenorhabditis elegans. This region of similarity, which lies outside the SH2 domain, also contains a pleckstrin homology domain. The presence of evolutionarily conserved domains indicates that GRB-7 is likely to perform a basic signaling function. The fact that GRB-7 and HER2 are both overexpressed and bound tightly together suggests that this basic signaling pathway is greatly amplified in certain breast cancers.
Insights
Growth factor receptor tyrosine kinases activate signaling pathways. GRB-7 protein binds to HER2, and both are overexpressed in breast cancer, suggesting amplified signaling in this disease.
Area of Science:
- Molecular Biology
- Oncology
- Signal Transduction
Background:
- SH2 domain proteins are crucial in signal transduction pathways initiated by growth factor receptor tyrosine kinases.
- Bacterial expression cloning identified novel SH2 domain proteins using a phosphorylated epidermal growth factor receptor C-terminus probe.
Purpose of the Study:
- To identify and characterize novel SH2 domain proteins involved in growth factor signaling.
- To investigate the role of the newly identified GRB-7 protein in breast cancer, particularly its relationship with HER2.
Main Methods:
- Bacterial expression cloning to identify SH2 domain proteins.
- Chromosomal mapping of GRB-7 in mice.
- Analysis of gene amplification and protein expression in breast cancer cell lines and tumors.
- Co-immunoprecipitation assays to assess protein binding interactions (GRB-7 with HER2 and SHC).
Main Results:
- GRB-7 was identified as a novel SH2 domain protein.
- GRB-7 maps to a region on mouse chromosome 11 syntenic to the human HER2 locus.
- GRB-7 is amplified and overexpressed concurrently with HER2 in breast cancer cell lines and tumors.
- GRB-7 binds tightly to tyrosine-phosphorylated HER2 and also binds to SHC.
- GRB-7 shares conserved domains (SH2 and pleckstrin homology) with a Caenorhabditis elegans gene, suggesting a fundamental signaling role.
Conclusions:
- GRB-7 is a novel SH2 domain protein implicated in growth factor receptor tyrosine kinase signaling.
- The co-amplification and overexpression of GRB-7 and HER2 in breast cancer suggest their joint contribution to amplified signaling pathways in tumorigenesis.
- GRB-7's conserved domains indicate a fundamental role in cellular signaling, which appears dysregulated in certain breast cancers.