PHOSPHATIDYLINOSITOL 8-KINASE-GAMMA ACTIVATES BRUTONS TYROSINE KINASEIN CONCERT WITH SRC FAMILY KINASES

Bruton's tyrosine kinase (Btk) is essential for normal B lymphocyte de velopment and function. The activity of Btk is partially regulated by transphosphorylation within its kinase domain by Src family kinases at residue Tyr-551 and subsequent autophosphorylation at Tyr-223. Activa tion correlates with Btk association with cellular membranes. Based on specific loss of function mutations, the Btk pleckstrin homology (PH) domain plays an essential role in this activation process. The Btk PH domain can bind in vitro to several lipid end products of the phospha tidylinositol 3-kinase (PI 3-kinase) family including phosphatidylinos itol 3,4,5-trisphosphate. Activation of Btk as monitored by elevation of phosphotyrosine content and a cellular transformation response was dramatically enhanced by coexpressing a weakly activated allel of Src (E378G) and the two subunits of PI 3-kinase-gamma. This activation cor relates with new sites of phosphorylation on Btk identified by two-dim ensional phosphopeptide mapping. Activation of Btk was dependent on th e catalytic activity of all three enzymes and an intact Btk PH domain and Src transphosphorylation site. These combined data define Btk as a downstream target of PI 3-kinase-gamma and Src family kinases.