The capacity of the host to fight intracellular patho- gens or tumours requires a combined response by the innate and adaptive immune systems, which recognize, respond and eliminate the offending cells. In particular, cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells identify and kill virus-infected or transformed cells. To activate CTLs, professional antigen-presenting cells (APCs) present, in a complex with MHC class I molecules, newly synthesized antigenic peptides or cross- present peptide antigens processed from endocytosed proteins derived from tumour cells or virus-infected cells (reviewed in REF. 1). NK cell cytotoxic activity is triggered by ligation of specific activating receptors and is inhibited by recognition of self MHC class I molecules. Thus recognition of self MHC class I molecules prevents killing of resting cells, whereas infected or tumour cells that have downregulated MHC class I expression, or stressed cells that upregulate expression of activating ligands, trigger NK cell activity (reviewed in REF. 2). CTLs and NK cells use both a perforin-dependent and a death receptor-dependent pathway to exert their cytotoxic function. These pathways have key functions in both immune surveillance and immune regulation. However, they are not functionally redundant. The death receptor pathway kills repeatedly stimulated T cells by ligation of FAS (also known as CD95), TNFRs (tumour necrosis factor receptors) or TRAILRs (TNF-related apoptosis inducing ligand receptors) with their cog- nate ligands (FASL, TNF and TRAIL, respectively) 3 . The FAS–FASL interaction participates to the control of T and B cell responses triggered by prolonged antigen stimulation. Ligand-induced oligomerization of death receptors leads to the formation of the death-inducing signalling complex that initiates caspase-dependent apoptosis. The perforin-dependent pathway involves the polarized secretion of cytotoxic granules that contain perforin and granzymes. Perforin is crucial for enabling access of granzymes into the target cell, which are involved in triggering caspase-dependent and caspase-independent cell death. Inherited deficiencies of both cytotoxic pathways have been described in humans and mice. Defects in either pathway are associated with dysregulation of lymphocyte homeostasis. Humans with inherited FAS or FASL deficiencies, as well as naturally occurring mouse mutants that lack FAS ( lpr and lpr cg mice) or FASL (gld mice), develop a lymphoproliferative syndrome, with an accumulation of non-malignant T and B cells, and autoimmunity 4 . The disease occurs owing to impaired antigen-induced cell death of repeatedly stimulated self- reactive T and B cells. This pathway will not be further discussed in this Review. By contrast, the phenotypic consequences of the genetic defects that affect effectors of the granule-dependent pathway provide evidence of an important role for this pathway in the regulation and/or termination of immune responses. In humans, defects in T and NK cell cytotoxic function through the granule-dependent pathway result in a severe and often fatal immunopathological condition known as haemo- phagocytic lymphohistiocytosis (HLH) 5,6 (TABLE 1). This *Institut National de la Santé et de la Recherche Médicale (INSERM), U768, Hôpital Necker Enfants Malades, 149 rue de Sèvres, 75015 Paris, France. ‡ Université Paris Descartes, Faculté de Médecine, 75015 Paris, France. § Centre d’Etude des Déficits Immunitaires, Assistance Publique-Hôpitaux de Paris, Hôpital Necker Enfants Malades, 75015 Paris, France. || AP-HP, Hôpital Necker Enfants Malades, Service d’Immunologie et d’Hématologie Pédiatrique, 75015 Paris, France. Correspondence to G.d.S.B. e-mail: genevieve.de-saint- basile@inserm.fr doi:10.1038/nri2803 Published online 16 July 2010 Molecular mechanisms of biogenesis and exocytosis of cytotoxic granules Geneviève de Saint Basile* ‡ § , Gaël Ménasché* ‡ and Alain Fischer* ‡|| Abstract | Cytotoxic T cells and natural killer cells are crucial for immune surveillance against virus-infected cells and tumour cells. Molecular studies of individuals with inherited defects that impair lymphocyte cytotoxic function have also highlighted the importance of cytotoxicity in the regulation and termination of immune responses. As discussed in this Review, characterization of these defects has contributed to our understanding of the key steps that are required for the maturation of cytotoxic granules and the secretion of their contents at the immunological synapse during target cell killing. This has revealed a marked similarity between cytotoxic granule exocytosis at the immunological synapse and synaptic vesicle exocytosis at the neurological synapse. We explore the possibility that comparison of these two kinetically and spatially regulated secretory pathways will provide clues to uncover additional effectors that regulate the cytotoxic function of lymphocytes. REVIEWS 568 | AUGUST 2010 | VOLUME 11 www.nature.com/reviews/immunol © 20 Macmillan Publishers Limited. All rights reserved 10