Lymphocyte activation is initiated by triggering of immunoreceptors (TCR, BCR, FcRs) by their natural ligands (eg, antigen/MHC in the case of T cells). Immediately thereafter, Src PTKs (Lck, Fyn, and Lyn) become activated by a mechanism that is still not completely understood, as discussed below, and in turn phosphorylate tandem tyrosine residues within specialized signaling motifs termed ITAMs (immunoreceptor tyrosine-based activation motifs; Billadeau and Leibson, this Perspective series, ref. 3). ITAMs are present in the cytoplasmic domains of the immunoreceptor-associated signal–transducing subunits such as CD3γ,-δ, and-ε, the TCRζ-chains, and the BCR-associated signaling molecules Igα and Igβ (CD79a and CD79b). Dually phosphorylated ITAMs provide the docking sites for the tandem SH2 domains of the Syk family PTKs ZAP-70 (in T cells) and Syk (in B cells), which are thereby targeted to the cell membrane and subsequently become activated through phosphorylation by Src PTKs. In T cells, one major substrate for ZAP-70 is LAT, which recruits several key signaling molecules to the plasma membrane after phosphorylation (4). For example, the three C-terminal TBSMs of LAT (Y171, Y191, and Y226) can mediate binding of Grb2 and its associated nucleotide exchange factor SOS (5). Which of these three tyrosines actually represents the major binding site for Grb2 in vivo is as yet unclear. Two of these TBSMs (Y171 and Y191) can also facilitate binding of Gads (5), a CAP that associates constitutively with SLP-76. Binding of Gads to LAT therefore brings SLP-76 to the membrane (6, 7). Initially, it was thought that, by binding to LAT, SLP-76 might be brought into the vicinity of ZAP-70, thus allowing its phosphorylation (8). However, the most recent analysis of Gads-deficient mice suggests that phosphorylation of SLP-76 may be independent of binding to LAT (9). In any case, phosphorylated SLP-76 provides the binding site for the SH2 domain of the tyrosine kinase Itk, to generate a membrane-associated complex consisting of LAT, Gads, SLP-76, and Itk (10).

Recruitment of Itk into this complex occurs in parallel with the binding of another key effector molecule, PLCγ1, to one of the remaining TBSMs in LAT (5). Thus, PLCγ1 is also integrated into the Ca2+-initiation complex, where it becomes activated by dual phosphorylation, mediated by ZAP-70 and Itk (Figure 3; see also ref. 11). Activated PLCγ1 cleaves the membrane-associated phosphoinositide PIP2, thus generating the second messengers IP3 and DAG. IP3 causes mobilization of Ca2+ from intra-and extracellular stores, thus raising intracellular Ca2+ levels, whereas DAG represents a classical activator of conventional and novel protein kinase C (PKC) isotypes (Figure 2). Among the PKC family members that are expressed in T cells, PKCθ stands out, since it seems to exert nonredundant functions that are essential for T cell activation. For example, PKCθ is required for activation of the transcription factors NF-κB and AP-1 (12, 13). Moreover, PKCθ appears to localize selectively into the center of the mature immunological synapse (IS; see Dustin, this Perspective series, ref. 14) during antigen stimulation (15). The elucidation of the molecular events underlying targeting of PKCθ to the IS is of par-