A phosphorylation site in Bruton's tyrosine kinase selectively regulates B cell calcium signaling efficiency by altering phospholipase C-gamma activation

Proc Natl Acad Sci U S A. 2004 Sep 28;101(39):14180-5. doi: 10.1073/pnas.0405878101. Epub 2004 Sep 16.


Loss of function of Bruton's tyrosine kinase (Btk) causes X-linked agammaglobulinemia (XLA) in humans and X-linked immunodeficiency in mice (xid). By using MS analysis and phosphopeptide-specific antibodies, we identified a tyrosine phosphorylation site (Y617) near the carboxyl terminus of the Btk domain from Btk expressed in 293T as well as DT-40 cells. Y617 is conserved in all Tec family kinases except murine Tec. Replacement of Y617 with a negatively charged glutamic acid (E) suppressed Btk-mediated phospholipase Cgamma2 activation and calcium response in DT-40 cells, whereas Akt activation was not affected. The Btk Y617E mutant could partially restore conventional B cell development and proliferation in Btk(-)/Tec(-) mice but failed to rescue CD5(+) B-1 cell development and the TI-II immune response to 2,4,6,-trinitrophenyl-Ficoll. These data suggest that Y617 phosphorylation or a negative charge at this site may down-regulate the function of Btk by selectively suppressing the B cell calcium signaling pathway.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Agammaglobulinaemia Tyrosine Kinase
  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • B-Lymphocytes / cytology
  • B-Lymphocytes / immunology
  • B-Lymphocytes / metabolism*
  • Binding Sites
  • CD5 Antigens / immunology
  • Calcium Signaling / physiology*
  • Cell Line
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Enzyme Activation / physiology
  • Glutamic Acid / genetics
  • Glutamic Acid / metabolism
  • Humans
  • Immunoprecipitation / methods
  • Mice
  • Phospholipase C gamma
  • Phosphorylation
  • Protein Structure, Tertiary
  • Protein-Tyrosine Kinases / chemistry
  • Protein-Tyrosine Kinases / deficiency
  • Protein-Tyrosine Kinases / genetics
  • Protein-Tyrosine Kinases / physiology*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Trinitrobenzenes / chemistry
  • Trinitrobenzenes / immunology
  • Trinitrobenzenes / pharmacology
  • Type C Phospholipases / antagonists & inhibitors
  • Type C Phospholipases / metabolism*


  • CD5 Antigens
  • Recombinant Proteins
  • Trinitrobenzenes
  • Glutamic Acid
  • Tec protein-tyrosine kinase
  • Protein-Tyrosine Kinases
  • Agammaglobulinaemia Tyrosine Kinase
  • BTK protein, human
  • Btk protein, mouse
  • Type C Phospholipases
  • Phospholipase C gamma