IntroductionSignaling through the B-cell receptor (BCR) evokes multiple signaling pathways that are required for the development and maintenance of B cells. 1,2 BCR engagement results in increased tyrosine phosphorylation mediated by nonreceptor protein tyrosine kinases (PTKs) of the Src, Syk, and Tec families. 3 The tyrosine residues located within immunoreceptor tyrosine-based activation motifs (ITAMs) of the intracellular domains of Ig␣/Ig␤ are phosphorylated by Lyn, a Src family kinase, and/or Syk. 4 Then, activated PTKs phosphorylate several substrates including coreceptors and cytoplasmic adaptor proteins such as BLNK and BCAP. [5][6][7] These adaptor and coreceptor proteins are thought to recruit effector enzymes, for instance phospholipase C␥2 (PLC␥2) and phosphoinositide 3-kinase (PI3K), by protein-protein and proteinlipid interactions, which in turn is essential for B-cell development and activation.CD19 is a well-known coreceptor that positively modulates BCR signaling and is suggested to set a threshold of BCR signaling. 8 In fact, upon BCR stimulation, CD19 undergoes phosphorylation on multiple tyrosine residues and recruits several signaling molecules that further augment transmembrane signaling. 9-14 Among these multiple tyrosine phosphorylation sites, the importance of 2 YXXM motifs in the CD19 cytoplasmic region, recruitment sites to the p85␣ of phosphoinositide 3-kinase (PI3K), has been validated by experiments using transgenic mice. 12 However, given that B-cell developmental defects in p85␣ Ϫ/Ϫ mice are more severe than those in CD19 Ϫ/Ϫ mice, 15-18 this CD19-dependent activation mechanism seems not to fully account for the activation mode of PI3K in B cells. Indeed, the BCR-mediated Akt activation was inhibited in CD19-deficient B cells, but not completely ablated. 19 Together, these observations suggest that other signaling molecule(s) could partially neutralize the defective PI3K activation in the absence of CD19. In this regard, BCAP could be a potential candidate. 20 BCAP, an adaptor molecule highly expressed in B cells, possesses binding ability to the p85␣ through its 4 YXXM motifs. 20 Indeed, BCAP underwent tyrosine phosphorylation after BCR stimulation, thereby recruiting the p85␣ of PI3K. However, B cells from BCAP-deficient mice exhibited apparently normal Akt activation, suggesting again that other molecule(s), presumably CD19, could rescue the BCAP function, particularly in the BCAP single-knockout mice. 21 Here, by using CD19/BCAP doubleknockout mice, we show that BCAP and CD19 have overlapping functions in BCR-mediated PI3K activation, thereby contributing to generation of immature and mature B cells. Methods MiceCD19 Ϫ/Ϫ and BCAP Ϫ/Ϫ mice were described previously. 15,21 CD19 Ϫ/Ϫ BCAP Ϫ/Ϫ mice were obtained by crossing with CD19 Ϫ/Ϫ and BCAP Ϫ/Ϫ mice. Six-to 8-week-old mice were used for all experiments. All mouse protocols were approved by RIKEN Animal Research Committee. Antibodies and reagentsThe following monoclonal antibodies (mAbs) and reagents were purchased from BD Bio...

C]leucine by U343MGa cells is Na؉ -independent and inhibited by BCH (2-amino-2-norbornane carboxylic acid), and branched and aromatic amino acids, indicating that the LAT1 is expressed at the cell surface. Pulse chase labeling and surface labeling experiments of this cell line indicate that the protein synthesis of LAT1 and 4F2hc is slow, however, the heterodimeric complex assembled in the cells is very stable, and that the disulfide bond between the LAT1 and 4F2hc is not directly involved in the stability of the heterodimer.