Credit: V. Summersby/Macmillan Publishers Limited

Apoptosis and necrosis are considered to be distinct modes of cell death; however, apoptosis can progress to secondary necrosis if apoptotic cells are not efficiently removed by phagocytic cells. Secondary necrosis was thought to be unregulated and to occur through passive cell swelling. However, Rogers et al. now show that secondary necrosis is another example of programmed cell death and that it is triggered by apoptotic stimuli.

Apoptosis is executed by the activation of a cascade of proteolytic enzymes, the caspases. Pyroptosis — a form of programmed necrosis induced by microbial infection — is also activated by caspases (in this case inflammatory caspases), which are responsible for the proteolytic processing of gasdermin D (GSDMD). The amino-terminal portion of GSDMD then oligomerizes and forms pores in the plasma membrane and thereby mediates necrotic cell death. The authors found that another member of the gasdermin superfamily, DFNA5, is also a caspase target, but unlike GSDMD it is cleaved by caspase 3, the executioner caspase of the apoptotic pathway. Akin to GSDMD, this processing occurs at a single site (Asp270) and generates amino-terminal (DFNA5-N) and carboxy-terminal (DFNA5-C) portions of DFNA5.

When DFNA5-N was ectopically expressed in human embryonic kidney 293T (HEK293T) cells, which do not express endogenous DFNA5, the cells showed morphological and biochemical features characteristic of necrotic cells. DFNA5-N was shown to associate with the plasma membrane and computer modelling approaches indicated that it can form pores, which indicates that DFNA5-N generated through caspase 3 cleavage is an inducer of necrosis.

Expression of DFNA5 (but not the non-cleavable mutant) and concomitant induction of apoptosis in HEK293T cells generated DFNA5-N and induced secondary necrosis. The DFNA5-N fragment was also detected when apoptosis was induced in macrophages, which endogenously express DFNA5. These macrophages progressed to secondary necrosis, which was impaired in DFNA5-deficient cells, indicating that DFNA5 mediates secondary necrosis and that its proteolytic processing downstream of apoptotic stimuli is essential for this activity.

secondary necrosis is a regulated process

This work shows that DFNA5 is a substrate of caspase 3 and drives necrosis after apoptosis induction, revealing that secondary necrosis is a regulated process downstream of apoptotic cell death. How this mode of cell death and its regulation contribute to physiological and pathological processes remains to be determined.