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Transactivation of Abl by the Crk II adapter protein requires a PNAY sequence in the Crk C-terminal SH3 domain

Abstract

To gain a better understanding of how Crk II regulates the function of the Abl tyrosine kinase, we explored the function of the C-terminal linker and SH3 domain, a region of Crk II that is still poorly understood. Molecular modeling, tryptophan fluorescence, and covariation sequence alignment indicate that the Crk-SH3-C has a unique binding groove and RT loop not observed in typical SH3 domains. Based on these models, we made a series of mutations in the linker and in residues predicted to destabilize the putative binding pocket and RT loop. In Abl transactivation assays, Y222F and P225A mutations in the linker resulted in strong transactivation of Abl by Crk II. However, mutations predicted to be at the surface of the Crk SH3-C were not activators of Abl. Interestingly, combinations of activating mutations of Crk II with mutations in the highly conserved PNAY sequence in the SH3-C inactivated the activating mutations, suggesting that the SH3-C is necessary for activation. Our data provide insight into the role of highly conserved residues in the Crk-SH3-C, suggesting a mechanism for how the linker and the Crk-SH3-C function in the transactivation of the Abl tyrosine kinase.

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Acknowledgements

We thank Drs Cecile Bougeret, Brain Kay, Tom Muir, Stephan Feller, Sally Kornbluth, and Allan Davidson for insightful discussions and sharing unpublished data. This work was supported by a Public Health Service Award (NIH-GMS55760) to RBB and an NJCCC fellowship to Sukhwinder Singh.

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Correspondence to Raymond B Birge.

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Reichman, C., Singh, K., Liu, Y. et al. Transactivation of Abl by the Crk II adapter protein requires a PNAY sequence in the Crk C-terminal SH3 domain. Oncogene 24, 8187–8199 (2005). https://doi.org/10.1038/sj.onc.1208988

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Keywords

  • Crk
  • Abl
  • SH3 domain
  • atypical
  • covariation
  • protein modeling
  • kinase activation

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