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Constitutive phosphorylation of Shc proteins in human tumors
G Pelicci1, L Lanfrancone, A E Salcini
1Istituto di Medicina Interna e Scienze Oncologiche, University of Perugia, Italy.
Abstract:
The Shc gene encodes three overlapping proteins which all contain a carboxy-terminal SH2 domain. Shc proteins are ubiquitously expressed and are downstream targets and effectors of activated tyrosine kinases (TK). We investigated tyrosine-phosphorylation of Shc proteins in normal and transformed cells. In tumor cells with known TK gene alterations Shc proteins were constitutively phosphorylated and complexed with the activated TK. No constitutive Shc phosphorylation was found in primary cell cultures and normal tissues. In 14 of 27 tumor cell lines with no reported TK alterations, Shc proteins were constitutively phosphorylated and formed stable complexes with novel tyrosine-phosphorylated polypeptides. Ten distinct Shc-associated phosphoproteins were identified with molecular weights ranging from 30 to 200 kDa. In a subset of carcinoma cell lines, phosphorylated Shc proteins complexed with a p175 phosphoprotein that was identified as the constitutively activated EGFR. In one glioblastoma cell line, a Shc-associated p190 was identified as the activated PDGFR. In 13 of 14 acute leukemia samples phosphorylated Shc proteins were constitutively complexed with a p140 phosphoprotein. Some of the Shc-associated phosphoproteins (EGFR, PDGFR, erbB-2, Met, bcr-abl, H4-ret) bound both the Shc- and Grb2-SH2 domains in vitro; others (p175; p70-p80) only the Shc-SH2 domain and yet others (p140) only the Grb2-SH3 domains. These results indicate that Shc proteins are common substrates of constitutively activated TKs and that the analysis of Shc phosphorylation allow the identification of tumors with constitutive TK activation.
Insights
Shc proteins are common targets of activated tyrosine kinases (TKs). Analyzing Shc phosphorylation can identify tumors with constitutive TK activation, aiding in cancer diagnosis.
Area of Science:
- Molecular Biology
- Cancer Biology
- Signal Transduction
Background:
- Shc proteins, containing an SH2 domain, are downstream effectors of tyrosine kinases (TKs).
- Constitutive activation of TKs is implicated in various cancers.
Purpose of the Study:
- To investigate tyrosine-phosphorylation of Shc proteins in normal and transformed cells.
- To determine if Shc phosphorylation can serve as a marker for constitutive TK activation in tumors.
Main Methods:
- Analysis of Shc protein tyrosine-phosphorylation in tumor cell lines and primary tissues.
- Co-immunoprecipitation to identify Shc-associated phosphoproteins.
- In vitro binding assays with SH2 domains.
Main Results:
- Shc proteins were constitutively phosphorylated and complexed with activated TKs in tumor cells, but not normal tissues.
- Fourteen of 27 tumor cell lines without known TK alterations showed constitutive Shc phosphorylation.
- Ten distinct Shc-associated phosphoproteins were identified, including EGFR, PDGFR, and a p140 phosphoprotein in leukemia samples.
Conclusions:
- Shc proteins are common substrates of constitutively activated TKs.
- Shc phosphorylation analysis is a valuable tool for identifying tumors with constitutive TK activation.
- This approach can aid in the molecular classification and potential targeted therapy of cancers.