he specificity of T-cell recognition events is essential for protective immune responses and in order to avoid autoimmunity. Binding of the T-cell receptor (TCR) to peptide bound to major histocompatibility complex (MHC) molecules is an early step for specific T-cell activation. This engagement leads to activation of the tyrosine kinase Lck and phosphorylation of immunoreceptor tyrosinebased activation (ITAM) motifs present within the CD3 complex, which activates ZAP-70. This process leads to the activation of Ras and the mitogen-activated protein (MAP) kinases ERK1 and -2. TCR engagement also leads to the activation of phospholipase Cβ₯ (PLCβ₯) and a subsequent increase in intracellular free Ca 2Ο© , together with the activation of protein kinase C (PKC). These two pathways converge at the level of interleukin 2 (IL-2) production, this being a critical parameter for T-cell activation [1][2][3][4] .
Phosphorylation of the CD3β₯ chain leads to internalization of the TCR-CD3 complex and marks it for degradation in the lysosome [5][6][7] . Because only functionally triggered TCRs are internalized, the extent of TCR downregulation closely correlates with T-cell activation (see below). The role of phosphorylation of the TCR-CD3 complex in internalization, compared with more downstream events such as changes in the cytoskeleton, is not fully understood. Studies using mutant cell lines have shown that maximal TCR internalization is dependent on early events such as functional Lck and/or PKC (Refs 8, 9). Surprisingly, however, TCR downregulation is barely affected by blocking-agents such as genistein, herbimycin or tyrphostin 10,11 . Thus, the residual phosphorylation activity left in the presence of such agents is apparently sufficient to allow for normal TCR internalization, and TCR downregulation is only affected if early signalling molecules are completely blocked.
In addition to these early signals, other events, such as Rab5mediated endocytosis, are important for TCR downregulation 12 . Moreover, TCR internalization can be inhibited by cytochalasin D, which selectively inhibits rearrangement of the actin cytoskeleton (M.F. Bachmann, unpublished). Thus, full TCR downregulation is dependent upon both early and late signalling events.