The full oncogenic activity of Ret/ptc2 depends on tyrosine 539, a docking site for phospholipase Cγ

Maria Grazia Borrello, Luisella Alberti, Elena Arighi, Italia Bongarzone, Carlo Battistini, Alberto Bardelli, Barbara Pasini, Claudia Piutti, Maria Grazia Rizzetti, Piera Mondellini, Maria Teresa Radice, Marco A. Pierotti

Research output: Contribution to journalArticle

Abstract

RET/PTC oncogenes, generated by chromosomal rearrangements in papillary thyroid carcinomas, are constitutively activated versions of proto-RET, a gene coding for a receptor-type tyrosine kinase (TK) whose ligand is still unknown. RET/PTCs encode fusion proteins in which proto-RET TK and C-terminal domains are fused to different donor genes. The respective Ret/ptc oncoproteins display constitutive TK activity and tyrosine phosphorylation. We found that Ret/ptcs associate with and phosphorylate the SH2-containing transducer phospholipase Cγ (PLCγ). Two putative PLCγ docking sites, Tyr- 505 and Tyr-539, have been identified on Ret/ptc2 by competition experiments using phosphorylated peptides modelled on Ret sequence. Transfection experiments and biochemical analysis using Tyr→Phe mutants of Ret/ptc2 allowed us to rule out Tyr-505 and to identify Tyr-539 as a functional PLCγ docking site in vivo. Moreover, kinetic measurements showed that Tyr-539 is able to mediate high-affinity interaction with PLCγ. Mutation of Tyr-539 resulted in a drastically reduced oncogenic activity of Ret/ptc2 on NIH 3T3 cells (75 to 90% reduction) both in vitro and in vivo, which correlates with impaired ability of Ret/ptc2 to activate PLCγ. In conclusion, this paper demonstrates that Tyr-539 of Ret/ptc2 (Tyr-761 on the proto-RET product) is an essential docking site for the full transforming potential of the oncogene. In addition, the present data identify PLCγ as a downstream effector of Ret/ptcs and suggest that this transducing molecule could play a crucial role in neoplastic signalling triggered by Ret/ptc oncoproteins.

Original languageEnglish
Pages (from-to)2151-2163
Number of pages13
JournalMolecular and Cellular Biology
Volume16
Issue number5
Publication statusPublished - May 1996

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

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    Borrello, M. G., Alberti, L., Arighi, E., Bongarzone, I., Battistini, C., Bardelli, A., Pasini, B., Piutti, C., Rizzetti, M. G., Mondellini, P., Radice, M. T., & Pierotti, M. A. (1996). The full oncogenic activity of Ret/ptc2 depends on tyrosine 539, a docking site for phospholipase Cγ. Molecular and Cellular Biology, 16(5), 2151-2163.