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Crosstalk between the catalytic and regulatory domains allows bidirectional regulation of Src

Abstract

The catalytic activity of Src family tyrosine kinases is inhibited by intramolecular interactions between the regulatory SH3 and SH2 domains and the catalytic domain. In the inactive state, the critical αC-helix in the catalytic domain is positioned such that the formation of the Glu 310–Lys 295 salt bridge is precluded, Tyr 416 in the activation loop is unphosphorylated, and the SH2 and SH3 domains are unavailable for interactions with other proteins. We found that phosphorylation of the activation loop or mutation of the loop preceding the αC-helix activates Src and increases the accessibility of the SH3 domain for ligands. Interaction of the αC-helix with the activation loop is a central component of this regulatory system. Our data suggest a bidirectional regulation mechanism in which the regulatory domains inhibit Src activity, and Src activity controls the availability of the regulatory domains. By this mechanism, Src family kinases can be activated by proteins phosphorylating or changing the conformation of the catalytic domain. Once active, Src family kinases become less prone to regulation, implying a positive feedback loop on their activity.

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Figure 1: Structure of Src. General view of the topology of Tyr 527-phosphorylated, regulated, human c-Src lacking the unique domain15.
Figure 2: Activity of Src mutants in mammalian cells. a, Phosphotyrosine levels of cellular proteins after transient transfection of various Src alleles in HEK293 cells
Figure 3: Regulation of Src/Lck chimeras
Figure 4: Role of Tyr 416
Figure 5: Residues participating in the αC-helix/activation loop communication.

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Acknowledgements

We wish to thank R. Wierenga for support, H. Pluk for the HA-paxillin construct, M.J. Eck for the human Src coordinates, T. Erpel and S.A. Courtneidge for the original SH3 domain swap and suggestions, A. Nelsbach for the anti-phospho-Y416 antibodies, J.R. Engen for help with the figures, A. Nebreda, R. Klein, M. Way and members of the Superti-Furga laboratory for suggestions and critical reading of the manuscript. S.G. was supported by a fellowship from the EC, the EMBL and the Boncompagni-Ludovisi Foundation. A.W. was supported by an EC fellowship.

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Correspondence to Giulio Superti-Furga.

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Gonfloni, S., Weijland, A., Kretzschmar, J. et al. Crosstalk between the catalytic and regulatory domains allows bidirectional regulation of Src. Nat Struct Mol Biol 7, 281–286 (2000). https://doi.org/10.1038/74041

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