Apoptosis is a form of physiological cell death, characterized by chromatin condensation, cytoplasmic blebbing and DNA fragmentation, which often depends on RNA and protein synthesis by the dying cell. The c-myc proto-oncogene, usually implicated in cell transformation, differentiation and cell-cycle progression also has a central role in some forms of apoptosis. These opposing roles of myc in cell growth and death require that other gene products dictate the outcome of c-Myc expression on a cell. A candidate for such a modifying gene is bcl-2, whose product prolongs cell survival and blocks apoptosis in some systems. Here we demonstrate that Bcl-2 prevents apoptotic death induced by c-Myc, provide a mechanism whereby cells can express c-Myc without undergoing apoptosis, and give a possible explanation for the ability of Bcl-2 to synergize with c-Myc in cell transformation.

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Fas receptor-induced apoptosis plays critical roles in immune homeostasis. However, most of the signal transduction events distal to Fas ligation have not been elucidated. Here, we show that Ras is activated following ligation of Fas on lymphoid lines. The activation of Ras is a critical component of this apoptotic pathway, since inhibition of Ras by neutralizing antibody or a dominant-negative Ras mutant interfered with Fas-induced apoptosis. Furthermore, ligation of Fas also resulted in stimulation of the sphingomyelin signalling pathway to produce ceramides, which, in turn, are capable of inducing both Ras activation and apoptosis. This suggests that ceramides acts as second messengers in Fas signaling via Ras. Thus, ligation of the Fas molecule on lymphocyte lines induces activation of Ras via the action of ceramide, and this activation is necessary, but not sufficient, for subsequent apoptosis.

Immature T cells and some T cell hybridomas undergo apoptotic cell death when activated through the T cell receptor complex, a phenomenon that is probably related to antigen induced negative selection of developing T cells. This activation-induced apoptosis depends on active protein and RNA synthesis in the dying cells, although none of the genes required for this process have previously been identified. Antisense oligonucleotides corresponding to c-myc block the constitutive expression of c-Myc protein in T cell hybridomas and interfere with all aspects of activation-induced apoptosis without affecting lymphokine production in these cells. These data indicate that c-myc expression is a necessary component of activation-induced apoptosis.

Mast cells derived from Bruton's tyrosine kinase (Btk)-defective xid or btk null mice showed greater expansion in culture containing interleukin-3 (IL-3) than those from wild-type (wt) mice. Although the proliferative response to IL-3 was not significantly different between the wt and xid mast cells, xid and btk null mast cells died by apoptosis more slowly than their wt counterparts upon IL-3 deprivation. Consistent with these findings, the apoptosis-linked c-Jun N-terminal kinase͞stress-activated protein kinase (JNK) activity was compromised in these btk-mutated cells upon FcRI crosslinking or upon stimulation with IL-3 or with stem cell factor. p38 activity was less severely, but significantly, affected by btk mutation, whereas extracellular signal-regulated kinases were not affected by the same mutation. Btk-mediated regulation of apoptosis and JNK activity was confirmed by reconstitution of btk null mutant mast cells with the wt btk cDNA. Furthermore, growth factor withdrawal induced the activation and sustained activity of JNK in wt mast cells, while JNK activity was consistently lower in btk-mutated mast cells. These results support the notion that Btk regulates apoptosis through the JNK activation.Bruton's tyrosine kinase (Btk) is a cytoplasmic proteintyrosine kinase of the recently recognized Tec family (1-3). These protein-tyrosine kinases consist, from the N to C termini, of pleckstrin homology, Tec homology, Src homology 3 (SH3), SH2, and SH1 (ϭkinase) domains. Various defects in the btk gene cause X-linked agammaglobulinemia in humans (2, 3) and the Arg to Cys mutation at position 28 results in X-linked immunodeficiency (xid) in mice (4, 5). These diseases and the phenotype of btk gene knockout mice (6, 7) indicate an essential role of Btk in the development and functions of B lymphocytes. Our previous study showed that Btk is tyrosinephosphorylated and enzymatically activated upon crosslinking of FcRI, suggesting a role of Btk in mast cell activation (8). Recent studies have made significant progresses in our understanding of how Btk activity is regulated. Thus, Btk was shown to be activated by phosphorylation of Tyr-551 by a Src family protein-tyrosine kinase, Lyn (9, 10). Activated Btk in turn autophosphorylates Tyr-223 in the SH3 domain (11). On the other hand, Btk is negatively regulated by protein kinase C in mast cells (12). Btk is also implicated in the regulation of intracellular Ca 2ϩ concentrations through direct or indirect phosphorylation of phospholipase C␥2 in chicken B lymphoma cells (13). However, the mechanisms by which Btk promotes normal functions in B cells and other cell types are largely unknown. In the present study, we provide evidence that Btk regulates the activity of c-Jun N terminal kinase or stressactivated protein kinase (JNK) and the apoptotic process upon growth factor deprivation of mast cells.