A phosphorylation site in Bruton's tyrosine kinase selectively regulates B cell calcium signaling efficiency by altering phospholipase C-gamma activation.
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.
Guo
Shuling
S
Department of Microbiology, Immunology, and Molecular Genetics and Howard Hughes Medical Institute, University of California, Los Angeles, CA 90095, USA.
Ferl
Gregory Z
GZ
Deora
Rajendar
R
Riedinger
Mireille
M
Yin
Sheng
S
Kerwin
James L
JL
Loo
Joseph A
JA
Witte
Owen N
ON
D000077329:Agammaglobulinaemia Tyrosine Kinase; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D001665:Binding Sites; D018956:CD5 Antigens / Q000276:immunology; D020013:Calcium Signaling / Q000502:physiology*; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004789:Enzyme Activation / Q000502:physiology; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D051379:Mice; D051966:Phospholipase C gamma; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014301:Trinitrobenzenes / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D018956:CD5 Antigens; D011994:Recombinant Proteins; D014301:Trinitrobenzenes; D018698:Glutamic Acid; C089848:Tec protein-tyrosine kinase; D011505:Protein-Tyrosine Kinases; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625949:BTK protein, human; C000625951:Btk protein, mouse; D010738:Type C Phospholipases; D051966:Phospholipase C gamma