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Key role of Shc signaling in the transforming pathway triggered by Ret/ptc2 oncoprotein
E Mercalli1, S Ghizzoni, E Arighi
1Department of Experimental Oncology, Research Unit #3, Istituto Nazionale Tumori, Via G. Venezian, 1 20133 Milan, Italy.
Abstract:
The RET/PTC oncogenes, generated by chromosomal rearrangements in papillary thyroid carcinomas, are constitutively activated versions of protoRET, a gene encoding two protein isoforms of a transmembrane tyrosine kinase receptor. By using Ret/ptc2 short isoform (iso9), we have previously demonstrated that Tyr586 (Tyr1062 of protoRet) is the docking site for both the PTB and the SH2 domains of Shc. To determine the relevance of this interaction for the transforming activity of Ret/ptc oncogenes, we have generated and characterized novel Ret/ptc mutants unable to activate Shc: Ret/ptc2 long isoform (iso51)-Y586F and both isoforms of Ret/ptc2-N583A. These mutants neither activate Shc nor transform NIH3T3 cells. Since Tyr1062 shows features of a multifunctional docking site, we have used a Shc mutant (Shc Y317F) to directly assess Shc role. We have demonstrated that in our cell system Shc Y317F behaves like a dominant interfering mutant on the activation of the Grb2-Sos pathway by endogenous Shc triggered by Ret/ptc2. A strong reduction of the transforming activity of Ret/ptc2 in presence of this mutant was also demonstrated. Our data suggest that Shc activation play a key role in the transforming pathways triggered by Ret/ptc oncoproteins. Moreover, we have shown that coexpression of the Shc-Y317F mutant with Ret/ptc2 specifically causes apoptosis, and that the surviving cells lose the long-term expression of one of the two genes.
Insights
RET/PTC oncogenes drive papillary thyroid cancer by activating the Shc protein. Mutating key sites on RET/PTC or Shc blocks transformation and induces apoptosis, highlighting Shc
Area of Science:
- Oncology
- Molecular Biology
- Cell Signaling
Background:
- RET/PTC oncogenes arise from chromosomal rearrangements in papillary thyroid carcinomas.
- These oncogenes are constitutively activated forms of protoRET, encoding a transmembrane tyrosine kinase receptor.
- The protoRET receptor has two protein isoforms.
Purpose of the Study:
- To investigate the role of Shc protein activation in the transforming activity of RET/PTC oncogenes.
- To characterize novel RET/PTC mutants unable to activate Shc.
- To assess the specific role of Shc in RET/PTC-mediated cellular transformation and signaling.
Main Methods:
- Generation and characterization of RET/PTC mutants (Ret/ptc2 iso51-Y586F, Ret/ptc2-N583A).
- Assessment of Shc activation by RET/PTC mutants in NIH3T3 cells.
- Utilizing a dominant interfering Shc mutant (Shc Y317F) to probe Shc function.
- Analysis of the Grb2-Sos pathway activation.
- Evaluation of cellular transformation and apoptosis induction.
Main Results:
- Novel RET/PTC mutants (Y586F, N583A) failed to activate Shc and transform NIH3T3 cells.
- The Shc Y317F mutant interfered with Shc activation of the Grb2-Sos pathway by Ret/ptc2.
- The Shc Y317F mutant significantly reduced the transforming activity of Ret/ptc2.
- Co-expression of Shc-Y317F and Ret/ptc2 induced apoptosis in cells.
- Surviving cells exhibited loss of long-term expression of one of the two genes.
Conclusions:
- Shc activation is crucial for the transforming pathways mediated by RET/PTC oncoproteins.
- Targeting Shc signaling represents a potential therapeutic strategy for RET/PTC-driven cancers.
- Specific RET/PTC-Shc interactions can lead to apoptosis, suggesting a complex regulatory mechanism.