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Identification of Shc docking site on Ret tyrosine kinase
E Arighi1, L Alberti, F Torriti
1Division of Experimental Oncology A, Istituto Nazionale Tumori, Milan, Italy.
Abstract:
The RET proto-oncogene encodes two isoforms of a receptor type tyrosine kinase which plays a role in neural crest and kidney development. Distinct germ-line mutations of RET have been associated with the inherited cancer syndromes MEN2A, MEN2B and FMTC as well as with the congenital disorder Hirschsprung disease (HSCR), whereas somatic rearrangements (RET/PTCs) have been frequently detected in the papillary thyroid carcinoma. Despite these findings, suggesting a relevant role for RET product in development and neoplastic processes, little is known about the signalling triggered by this receptor. In this study, we have demonstrated that the transducing adaptor molecule Shc is recruited and activated by both Ret isoforms and by the rearranged cytoplasmatic Ret/ptc2 oncoproteins as well as by the membrane bound receptor activated by MEN2A or by MEN2B associated mutations. Moreover, our analysis has identified the Ret tyrosine residue and the Shc domains involved in the interaction. In fact, here we show that both the phosphotyrosine binding domains of Shc, PTB and SH2, interact with Ret/ptc2 in vitro. However, PTB domain binds 20 folds higher amount of Ret/ptc2 than SH2. The putative binding site for either SH2 and PTB domains has been identified as Tyr586 of Ret/ptc2 (Tyr1062 on proto-Ret). In keeping with this finding, by using RET/PTC2-Y586F mutant, we have demonstrated that this tyrosine residue, the last amino acid but one before the divergence of the two Ret isoforms, is the docking site for Shc.
Insights
The RET receptor tyrosine kinase interacts with the Shc adaptor protein, crucial for its activation. This study identifies the specific RET tyrosine residue (Tyr586) and Shc domains (PTB and SH2) involved in this signaling pathway.
Area of Science:
- Oncology
- Molecular Biology
- Developmental Biology
Background:
- The RET proto-oncogene encodes receptor tyrosine kinases vital for neural crest and kidney development.
- RET mutations are linked to inherited cancers (MEN2A, MEN2B, FMTC) and Hirschsprung disease (HSCR).
- Somatic RET rearrangements (RET/PTCs) are common in papillary thyroid carcinoma, but RET signaling remains poorly understood.
Purpose of the Study:
- To investigate the signaling pathways triggered by RET receptor tyrosine kinases.
- To determine if the Shc adaptor protein is involved in RET-mediated signaling.
- To identify the specific molecular interactions between RET and Shc.
Main Methods:
- Investigated the recruitment and activation of the Shc adaptor molecule by RET isoforms and rearranged RET/PTC2 oncoproteins.
- Analyzed interactions between Shc domains (PTB, SH2) and RET/PTC2 in vitro.
- Utilized a RET/PTC2-Y586F mutant to identify the critical tyrosine residue for Shc binding.
Main Results:
- Demonstrated that Shc is recruited and activated by both Ret isoforms and rearranged Ret/ptc2 oncoproteins.
- Identified Tyr586 of Ret/ptc2 (Tyr1062 on proto-Ret) as the specific binding site for Shc's PTB and SH2 domains.
- Showed that Shc's PTB domain binds Ret/ptc2 with significantly higher affinity than the SH2 domain.
Conclusions:
- The Shc adaptor protein is a key component of RET signaling pathways.
- Specific tyrosine residues and Shc domains mediate the interaction between RET and Shc.
- Understanding this interaction provides insights into RET's role in development and cancer.