RESUMO
Gasdermin D (GSDMD) is the executor of pyroptosis, which is important for host defence against pathogen infection. Following activation, caspase-mediated cleavage of GSDMD releases an amino-terminal fragment (GSDMD-NT), which oligomerizes and forms pores in the plasma membrane, leading to cell death and release of proinflammatory cytokines. The spatial and temporal regulation of this process in cells remains unclear. Here we identify GSDMD as a substrate for reversible S-palmitoylation on C192 during pyroptosis. The palmitoyl acyltransferase DHHC7 palmitoylates GSDMD to direct its cleavage by caspases. Subsequently, palmitoylation of GSDMD-NT promotes its translocation to the plasma membrane, where APT2 depalmitoylates GSDMD-NT to unmask the C192 residue and promote GSDMD-NT oligomerization. Perturbation of either palmitoylation or depalmitoylation suppresses pyroptosis, leading to increased survival of mice with lipopolysaccharide-induced lethal septic shock and increased sensitivity to bacterial infection. Our findings reveal a model through which a palmitoylation-depalmitoylation relay spatiotemporally controls GSDMD activation during pyroptosis.
Assuntos
Aciltransferases , Gasderminas , Lipoilação , Proteínas de Ligação a Fosfato , Piroptose , Animais , Humanos , Camundongos , Acetiltransferases , Aciltransferases/metabolismo , Aciltransferases/genética , Caspases/metabolismo , Membrana Celular/metabolismo , Células HEK293 , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Lipopolissacarídeos/farmacologia , Camundongos Endogâmicos C57BL , Proteínas de Neoplasias/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Ligação a Fosfato/metabolismo , Proteínas de Ligação a Fosfato/genética , Piroptose/genética , Choque Séptico/metabolismo , Choque Séptico/patologia , Choque Séptico/genéticaRESUMO
Vesicle associated membrane protein 2 (VAMP2/synaptobrevin2), a core SNARE protein residing on synaptic vesicles (SVs), forms helix bundles with syntaxin-1 and SNAP25 for the SNARE assembly. Prior to the SNARE assembly, the structure of VAMP2 is unclear. Here, by using in-cell NMR spectroscopy, we describe the dynamic membrane association of VAMP2 SNARE motif in mammalian cells, and the structural change of VAMP2 upon the change of intracellular lipid environment. We analyze the lipid compositions of the SV membrane by mass-spectrometry-based lipidomic profiling, and further reveal that VAMP2 forms distinctive conformations in different membrane regions. In contrast to the non-raft region, the membrane region of cholesterol-rich lipid raft markedly weakens the membrane association of VAMP2 SNARE motif, which releases the SNARE motif and facilitates the SNARE assembly. Our work reveals the regulation of different membrane regions on VAMP2 structure and sheds light on the spatial regulation of SNARE assembly.