Signaling through the EBV/C3d receptor (CR2, CD21) in human B lymphocytes:Activation of phosphatidylinositol 3-kinase via a CD19-independent pathway

We herein analyzed the regulation of phosphatidylinositol 3-kinase (PI 3-ki nase) activity by CR2 activated on B lymphocyte cell surface. We demonstrat ed that CR2 activation triggered in vivo PI 3-kinase activity and interacti on of PI 3-kinase p85 subunit with a tyrosine-phosphorylated p95 component. The specificity of PI 3-kinase activity was controlled using wortmannin an d LY294002, CR2 activation did not trigger tyrosine phosphorylation of PI 3 -kinase p85 subunit, but induced direct interaction of tyrosine phosphoryla ted p95 with the Src homology 2 domain of p85 subunit, as shown using gluta thione-S-transferase fusion proteins. Despite identical molecular masses, i mmunoblotting analysis demonstrated that tyrosine-phosphorylated p95 that i nteracted in vivo and in vitro with p85 was neither CD19, the 95-kDa proto- oncogene vav, nor Gab1 (a 95-kDa adaptor molecule). Furthermore, p95 tyrosi ne phosphoprotein also expressed in K562A cells (CR2(+) CD19(-) cells) inte racted with Src homology 2 domain of PI 3-kinase p85 subunit after CR2 acti vation. Activated CR2 did not interact directly with p85 subunit or tyrosin e-phosphorylated p95, This suggests the presence of an intermediate molecul e between activated CR2 and tyrosine-phosphorylated p95, which may be 3BP2. In addition, in contrast to CD19 activation, CR2 activation did not trigge r interaction of CD19 or Vav with PI 3-kinase p85 subunit or coprecipitatio n of PI 3-kinase activity with CD19, Together, these data clearly demonstra ted that CR2 activation triggered in vivo PI 3-kinase activation through a pathway distinct from that triggered through CD19 activation.