RESUMO
In concert with the TCR, CD28 promotes T cell survival by regulating the expression of the antiapoptotic protein Bcl-x(L). The mechanism by which CD28 mediates the induction of Bcl-x(L) remains unknown. We show that although signaling through the TCR is sufficient to stimulate transcription of Bcl-x(L) mRNA, CD28, by activating PI3K and mammalian target of rapamycin, provides a critical signal that regulates the translation of Bcl-x(L) transcripts. We observe that CD28 induced 4E-binding protein-1 phosphorylation, an inhibitor of the translational machinery, and that CD28 costimulation directly augmented the translation of a Bcl-x(L) 5'-untranslated region reporter construct. Lastly, costimulation by CD28 shifted the distribution of Bcl-x(L) mRNA transcripts from the pretranslation complex to the translationally active polyribosomes. These results demonstrate that CD28 relieves the translational inhibition of Bcl-x(L) in a PI3K/mammalian target of rapamycin-dependent manner.
Assuntos
Antígenos CD28/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Biossíntese de Proteínas/imunologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Linfócitos T/imunologia , Animais , Northern Blotting , Antígenos CD28/genética , Antígenos CD28/imunologia , Complexo CD3/imunologia , Complexo CD3/metabolismo , Morte Celular/fisiologia , Células Cultivadas , Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Humanos , Imunossupressores/farmacologia , Interleucina-2/biossíntese , Interleucina-2/imunologia , Células Jurkat , Camundongos , Camundongos Transgênicos , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases/imunologia , Biossíntese de Proteínas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/imunologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sirolimo/farmacologia , Linfócitos T/metabolismo , Transfecção , Proteína bcl-XRESUMO
Activation-induced cell death (AICD) is a phenomenon in which activated T cells undergo apoptosis upon restimulation. We are studying a form of AICD that can occur before cells become competent to die by Fas (hence "early" AICD) and which depends on the presence of perforin. Previous studies indicate that it does not occur through granule exocytosis but via some endogenous pathway. We here investigate a possible role for caspases. Caspase 3(-/-) cells were protected, suggesting a role for caspase 3 in early AICD. After recrosslinking, caspase 3 activity could be detected in cell lysates between 3 and 12 h, and CD8(+) T cells became annexin V-positive between 15 and 18 h. Blocking anti-Fas ligand antibody failed to inhibit death, and no processing of either caspase 8 or caspase 9 was detected in recrosslinked cells. Furthermore, T cells lacking functional caspase 9 continued to die in early AICD. Thus, perforin-dependent early AICD appears to require activation of caspase 3, but not caspases 8 or 9. As perforin has no intrinsic catalytic abilities, we propose that it releases some endogenous activity that can activate caspase 3.
