Volume 7 Supplement 1

12th Joint Meeting of the Signal Transduction Society (STS). Signal Transduction: Receptors, Mediators and Genes

Open Access

Elucidation of the SLP-65 phosphorylation state in activated B lymphocytes

  • T Oellerich1,
  • M Gronborg2,
  • K Neumann1,
  • HH Hsiao2,
  • H Urlaub2 and
  • J Wienands1
Cell Communication and Signaling20097(Suppl 1):A21

DOI: 10.1186/1478-811X-7-S1-A21

Published: 26 February 2009

The SH2 domain-containing leukocyte adaptor protein of 65 kDa (SLP-65) is a central effector for signaling downstream of the B-cell antigen receptor (BCR). Upon phosphorylation on serine/threonine and tyrosine residues, SLP-65 nucleates the formation of multimolecular protein complexes to integrate numerous BCR-signaling events. We have now qualitatively and quantitatively identified phospho acceptor sites of SLP-65 by applying state of the art mass spectrometry. SLP-65 turned out to possess a plethora of phospho acceptor sites. In fact, it turned out to be one of the most phosphorylated proteins described so far. Moreover, by applying stable isotope labelling of amino acids in cell culture (SILAC) we identified several acceptor sites whose phosphorylation kinetic is differentially regulated upon BCR-stimulation. The functional relevance of some of these sites was subsequently analyzed by mutational analysis of SLP-65 in SLP-65-deficient DT40 B cells. In contrast to the described role of SLP-65 tyrosine phosphorylation for the initiation of Ca2+-signaling, serine/threonine phosphorylation of SLP-65 turned out to be a key regulator for BCR-dependent MAP-kinase activation and AP-1 regulated gene transcription. Collectively our data explain several of the SLP-65 controlled biological responses elucidated by genetic means and further support the role of SLP-65 as the key integrator of BCR-signaling. In general (phospho)proteomics combined with reconstitution experiments in gene-targeted DT40 cells proves to be a powerful strategy to uncover post translational modifications and their biological relevance in cell signaling.

Authors’ Affiliations

(1)
Institute of Cellular and Molecular Immunology, Georg-August University
(2)
Max Planck Institute for Biophysical Chemistry, Bioanalytical Mass Spectrometry Group

Copyright

© Oellerich et al; licensee BioMed Central Ltd. 2009

This article is published under license to BioMed Central Ltd.

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