Multivesicular bodies (MVBs) are endocytic compartments that contain intraluminal vesicles formed by inward budding from the limiting membrane of endosomes. In T lymphocytes, these vesicles contain pro-apoptotic Fas ligand (FasL), which may be secreted as 'lethal exosomes' upon fusion of MVBs with the plasma membrane. Diacylglycerol kinase a (DGKa) regulate the secretion of exosomes, but it is unclear how this control is mediated. T-lymphocyte activation increases the number of MVBs that contain FasL. DGKa is recruited to MVBs and to exosomes in which it has a double function. DGKa kinase activity exerts a negative role in the formation of mature MVBs, as we demonstrate by the use of an inhibitor. Downmodulation of DGKa protein resulted in inhibition of both the polarisation of MVBs towards immune synapse and exosome secretion. The subcellular location of DGKa together with its complex role in the formation and polarised traffic of MVBs support the notion that DGKa is a key regulator of the polarised secretion of exosomes. 2 FasL induces crosslinking of the Fas death receptor on the target cell and apoptosis.3 In CTLs, FasL is located at the limiting membrane of secretory lysosomes containing perforin/granzyme with MVB structure.2 Upon T-cell receptor activation (TCR), MVBs undergo fusion with the PM, and relocalisation of FasL to the PM occurs. 2In addition to this transport of FasL, another mechanism for the delivery of FasL co-exists in CTLs. 4 This mechanism is because of the fact that FasL can be sorted from the limiting membrane of the MVBs to the ILVs by inward budding. 4,5 Upon cell activation, the fusion of preexisting MVBs with the PM results in the release of exosomes containing FasL. These authors contributed equally to this work.

Fas ligand (FasL) mediates both apoptotic and inflammatory responses in the immune system. FasL function critically depends on the different forms of FasL; soluble Fas ligand lacking the transmembrane and cytoplasmic domains is a poor mediator of apoptosis, whereas fulllength, membrane-associated FasL (mFasL) is pro-apoptotic. mFasL can be released from T lymphocytes, via the secretion of mFasL-bearing exosomes. mFasL in exosomes retains its activity in triggering Fas-dependent apoptosis, providing an alternative mechanism of cell death that does not necessarily imply cell-to-cell contact. Diacylglycerol kinase ␣ (DGK␣), a diacylglycerol (DAG)-consuming enzyme, is involved in the attenuation of DAG-derived responses initiated at the plasma membrane that lead to T lymphocyte activation. Here we studied the role of DGK␣ on activation-induced cell death on a T cell line and primary T lymphoblasts. The inhibition of DGK␣ increases the secretion of lethal exosomes bearing mFas ligand and subsequent apoptosis. On the contrary, the overactivation of the DGK␣ pathway inhibits exosome secretion and subsequent apoptosis. DGK␣ was found associated with the trans-Golgi network and late endosomal compartments. Our results support the hypothesis that the DGK␣ effect on apoptosis occurs via the regulation of the release of lethal exosomes by the exocytic pathway, and point out that the spatial orchestration of the different pools of DAG (plasma membrane and Golgi membranes) by DGK␣ is crucial for the control of cell activation and also for the regulation of the secretion of lethal exosomes, which in turn controls cell death.