Editors' ChoiceImmunology

Pore-forming death signal

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Science Signaling  19 Jul 2016:
Vol. 9, Issue 437, pp. ec163
DOI: 10.1126/scisignal.aah5655

Activation of the inflammatory caspases by pathogen-derived molecules or host danger signals results in pyroptosis, a lytic form of cell death that stimulates inflammation. Pyroptosis is triggered by the caspase-dependent cleavage of gasdermin D (GSDMD) such that its N-terminal domain (GSDMD-N), which has pyroptotic activity, is separated from the inhibitory C-terminal domain (GSDMD-C) of the protein. Three studies have identified the mechanism underlying the pyroptotic activity of GSDMD-N.

Ding et al. found that GSDMD-N, but not GSDMD-C, bound to phospholipid-containing liposomes. Fluorescence microscopy showed that tagged GSDMD-N was recruited to the plasma membrane of HeLa cells during pyroptosis. In liposomes, GSDMD-N molecules oligomerized and formed pores through which the liposome contents leaked. Solving the crystal structure of GSDMD identified the interaction points between the N- and C-terminal domains, and mutation of some of these residues resulted in spontaneous pyroptotic activity of the whole protein.

Liu et al. used electron microscopy to show that GSDMD-N molecules oligomerized and formed pores in liposomes containing phosphatidylserine, which is usually found in the inner leaflets of plasma membranes, or cardiolipin, which is found in both the inner and outer leaflets of bacterial membranes and in mitochondrial membranes. Mutation of four conserved positively charged residues in GSDMD-N blocked its ability to bind to membranes, oligomerize, and form pores. Release of GSDMD-N from cells that had undergone pyroptosis did not kill bystander cells, likely as a result of its phospholipid-binding preferences. However, extracellular GSDMD-N killed bacteria in vitro in a dose-dependent manner, raising the possibility that it might also have intracellular bactericidal activity.

Aglietti et al. cleaved GSDMD in vitro with constitutively active caspase-11 and found that the N-terminal product specifically interacted with liposomes with a plasma membrane-like composition or a mitochondrial-like composition and triggered the release of calcium from calcium-loaded liposomes. The GSDMD-N also cofractionated with membranes when produced by caspase-11 activation in macrophages, and transfection of HEK293 cells with GSDMD-N resulted in cell death. Electron microscopy of cleaved GSDMD in the presence of liposomes revealed ring-like structures. Analysis of the protein extracted from the liposomes suggested that the rings are comprised of ~24 monomers of GSDMD-N.

Together, these data indicate that after cleavage of GSDMD by inflammatory caspases, its N-terminal domain binds to the plasma membrane, oligomerizes, and forms pores that kill cells by pyroptosis. Furthermore, GSDMD-N may also contribute to killing of intracellular bacteria, which would limit the spread of infection following cell death by pyroptosis.

J. Ding, K. Wang, W. Liu, Y. She, Q. Sun, J. Shi, H. Sun, D.-C. Wang, F. Shao, Pore-forming activity and structural autoinhibition of the gasdermin family. Nature 535, 111–116 (2016). [PubMed]

X. Liu, Z. Zhang, J. Ruan, Y. Pan, V. G. Magupalli, H. Wu, J. Lieberman, Inflammasome-activated gasdermin D causes pyroptosis by forming membrane pores. Nature 535, 153–158 (2016). [PubMed]

R. A. Aglietti, A. Estevez, A. Gupta, M. Gonzalez Ramirez, P. S. Liu, N. Kayagaki, C. Ciferri, V. M. Dixit, E. C. Dueber, GsdmD p30 elicited by caspase-11 during pyroptosis forms pores in membranes. Proc. Natl. Acad. Sci. U.S.A. 113, 7858–7863 (2016). [PubMed]

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