The Caspase-1 Digestome Identifies the Glycolysis Pathway as a Target during Infection and Septic Shock

Abstract

Caspase-1 is an essential effector of inflammation, pyroptosis, and septic shock. Few caspase-1 substrates have been identified to date, and these substrates do not account for its wide range of actions. To understand the function of caspase-1, we initiated the systematic identification of its cellular substrates. Using the diagonal gel proteomic approach, we identified 41 proteins that are directly cleaved by caspase-1. Among these were chaperones, cytoskeletal and translation machinery proteins, and proteins involved in immunity. A series of unexpected proteins along the glycolysis pathway were also identified, including aldolase, triose-phosphate isomerase, glyceraldehyde-3-phosphate dehydrogenase, ␣-enolase, and pyruvate kinase. With the exception of the latter, the identified glycolysis enzymes were specifically cleaved in vitro by recombinant caspase-1, but not caspase-3. The enzymatic activity of wild-type glyceraldehyde-3-phosphate dehydrogenase, but not a non-cleavable mutant, was dampened by caspase-1 processing. In vivo, stimuli that fully activated caspase-1, including Salmonella typhimurium infection and septic shock, caused a pronounced processing of these proteins in the macrophage and diaphragm muscle, respectively. Notably, these stimuli inhibited glycolysis in wildtype cells compared with caspase-1-deficient cells. The systematic characterization of caspase-1 substrates identifies the glycolysis pathway as a caspase-1 target and provides new insights into its function during pyroptosis and septic shock.Caspases are aspartate-specific cysteine proteases known for their function in regulating programmed cell death and inflammation. Phylogenetically, they are subdivided into the CED3-related enzymes that initiate and execute cell death and the caspase-1-related proteins that process and mature cytokines, viz. IL-1␤, 2 IL-18, and IL-33. Apoptotic caspases execute cell death through the restricted cleavage of key cellular proteins required to maintain cell viability, resulting in the morphological changes observed during apoptosis, including membrane blebbing, nuclear condensation, and cytoskeletal dismantling (1). Caspase-1 is essential during inflammation because of its role in the activation of cytokine signaling pathways. With the exception of its cytokine substrates, very little is known regarding the spectrum of cellular proteins it targets upon full activation. Similar to other caspases, caspase-1 is found in cells as an inactive precursor and is activated in response to inflammatory triggers, including pathogen-derived molecules, as well as danger signals released from infected or dying cells (2). Caspase-1 activation is achieved in a macromolecular complex known as the inflammasome through its recruitment to a scaffolding molecule generally via the adaptor ASC (3). Scaffolding molecules that activate caspase-1 within the inflammasome belong to the cytosolic Nod-like family of pathogen recognition receptors and include Nalp1-3, Ipaf, and Naip5 (4). More recently, a distinct caspase-1 activat...

Inflammasome is a central player in the induction of obesity and insulin resistance

“…We postulate that caspase-1 downstream signaling contributes to the enhanced energy expenditure observed in HFD-fed Casp1 −/− animals. Indeed, using the diagonal gel proteomic approach, several proteins that are involved in energy metabolism were identified as caspase-1 substrates (31). Additional substrates of caspase-1 in adipose tissue may be identified by in silico analysis and may help to explain its contribution to the development of adipose tissue dysfunction during HFD-induced obesity.…”

Cell death in the host response to infection

“…Therefore, we propose that processing of substrates by caspase-1 is responsible for the phenotype of pyroptotic cells. 70 Caspase-1 also appears to be required for unconventional protein secretion. 71 The release of cytoplasmic contents is a highly inflammatory event, which underscores the distinct form and function of this programmed cell death.…”

“…99 The elucidation of the caspase-1 digestome identified additional substrates of caspase-1 that are thought to contribute to its effects during the host response to infections and in septic shock. 70 An additional layer of complexity can be attributed to redundancy in pathogeninduced host cell death. Although caspase-1 deficiency renders macrophages resistant to rapid pyroptotic cell death, these cells will nonetheless perish following infection due to the engagement of alternate death pathways.…”

Programmed cell death in the plant immune system

“…36 The precise mechanism by which caspase-1 leads to cell death has also been investigated by looking at caspase-1 substrates during infection and inflammation. The caspase-1 digestome includes chaperones, cytoskeletal maintenance components and proteins involved in energy metabolism, 37 as well as caspase-7, 38 which has been shown to be activated downstream of NLRC4 inflammasome during bacterial infection. 39 However, it is still not clear why pyroptotic cells die.…”