Mapping of p140Cap phosphorylation sites: the EPLYA and EGLYA motifs have a key role in tyrosine phosphorylation and Csk binding, and are substrates of the Abl kinase. - Université Grenoble Alpes Accéder directement au contenu
Article Dans Une Revue PLoS ONE Année : 2012

Mapping of p140Cap phosphorylation sites: the EPLYA and EGLYA motifs have a key role in tyrosine phosphorylation and Csk binding, and are substrates of the Abl kinase.

Simona Aramu
  • Fonction : Auteur
Nanaocha Sharma
  • Fonction : Auteur
Silvia Grasso
  • Fonction : Auteur
Isabella Russo
  • Fonction : Auteur
Ole N Jensen
  • Fonction : Auteur
Sara Cabodi
  • Fonction : Auteur
Emilia Turco
  • Fonction : Auteur
Paola Di Stefano
  • Fonction : Auteur
Paola Defilippi
  • Fonction : Auteur

Résumé

Protein phosphorylation tightly regulates specific binding of effector proteins that control many diverse biological functions of cells (e. g. signaling, migration and proliferation). p140Cap is an adaptor protein, specifically expressed in brain, testis and epithelial cells, that undergoes phosphorylation and tunes its interactions with other regulatory molecules via post-translation modification. In this work, using mass spectrometry, we found that p140Cap is in vivo phosphorylated on tyrosine (Y) within the peptide GEGLpYADPYGLLHEGR (from now on referred to as EGLYA) as well as on three serine residues. Consistently, EGLYA has the highest score of in silico prediction of p140Cap phosphorylation. To further investigate the p140Cap function, we performed site specific mutagenesis on tyrosines inserted in EGLYA and EPLYA, a second sequence with the same highest score of phosphorylation. The mutant protein, in which both EPLYA/EGLYA tyrosines were converted to phenylalanine, was no longer tyrosine phosphorylated, despite the presence of other tyrosine residues in p140Cap sequence. Moreover, this mutant lost its ability to bind the C-terminal Src kinase (Csk), previously shown to interact with p140Cap by Far Western analysis. In addition, we found that in vitro and in HEK-293 cells, the Abelson kinase is the major kinase involved in p140Cap tyrosine phosphorylation on the EPLYA and EGLYA sequences. Overall, these data represent an original attempt to in vivo characterise phosphorylated residues of p140Cap. Elucidating the function of p140Cap will provide novel insights into its biological activity not only in normal cells, but also in tumors.

Dates et versions

hal-01322391 , version 1 (27-05-2016)

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Citer

Daniele Repetto, Simona Aramu, Elisabetta Boeri Erba, Nanaocha Sharma, Silvia Grasso, et al.. Mapping of p140Cap phosphorylation sites: the EPLYA and EGLYA motifs have a key role in tyrosine phosphorylation and Csk binding, and are substrates of the Abl kinase.. PLoS ONE, 2012, 8 (1), pp.e54931. ⟨hal-01322391⟩
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