Active Caspase-3 Is Stored within Secretory Compartments of Viable Mast Cells

Abstract

Caspase-3 is a main executioner of apoptotic cell death. The general notion is that, in viable cells, caspase-3 is found as a cytosolic inactive proenzyme and that caspase-3 activation is largely confined to processes associated with cell death. In this study, we challenge this notion by showing that enzymatically active caspase-3 is stored in viable mast cells. The enzymatically active caspase-3 was undetectable in the cytosol of viable cells, but was recovered in subcellular fractions containing secretory granule-localized proteases. Moreover, active caspase-3 was rapidly released into the cytosolic compartment after permeabilization of the secretory granules. Using a cell-permeable substrate for caspase-3, the presence of active caspase-3–like activity in granule-like compartments close to the plasma membrane was demonstrated. Moreover, it was shown that mast cell activation caused release of the caspase-3 to the cell exterior. During the course of mast cell differentiation from bone marrow cells, procaspase-3 was present in cells of all stages of maturation. In contrast, active caspase-3 was undetectable in bone marrow precursor cells, but increased progressively during the process of mast cell maturation, its accumulation coinciding with that of a mast cell–specific secretory granule marker, mouse mast cell protease 6. Together, the current study suggests that active caspase-3 can be stored within secretory compartments of viable mast cells.

Extracellular TDP‐43 aggregates target MAPK/MAK/MRK overlapping kinase (MOK) and trigger caspase‐3/IL‐18 signaling in microglia

“…3). In addition, we observed an increase in cleaved caspase‐3 in the form of granules—similar to those previously described in viable mast cells (46)—as a result of TDP43 treatment (Fig. 3).…”

“…Furthermore, Hsp27 was found to dramatically suppress the number of caspase‐3 granules of TDP‐43–exposed cells and to slightly increase activated caspase‐3 total levels, which supports a role for caspase‐3 in IL‐18 alternative processing in microglia. Of interest, active caspase‐3 granule‐like compartments have been previously found in viable mast cells (46); therefore, the observed results with Hsp27 cotreatment may suggest an actual increase in caspase‐3 intracellular availability. If so, the incremented neurotoxic effects of TDP‐43–exposed microglia by Hsp27 cotreatment may be related to an exacerbated neuroinflammatory/neurotoxic profile that is linked to IL‐1β/‐18 and activated caspase‐3 (44, 67).…”

Secretory lysosomes of mouse mast cells store and exocytose active caspase‐3 in a strictly granzyme B dependent manner

“…The presence of active caspase‐3 in MC granules was recently reported . We extend this observation by employing WT and gzmB‐deficient mice to show that cleavage of pro‐caspase‐3 and its granular storage is strictly regulated by gzmB.…”

“…1E), suggesting that the processed form of caspase-3 represents fully operative caspase-3. Not explainable at the moment is the observation that supernatants of unstimulated BMMCs also contain caspase-3-like activity, which was also reported by Garcia-Faroldi et al [6]. Previously, we reported that gzmB is stored in secretory lysosomes of mouse MCs, both in vivo and in vitro [7].…”

Mefloquine, an anti‐malaria agent, causes reactive oxygen species‐dependent cell death in mast cells via a secretory granule‐mediated pathway

“…After washing, cells were resuspended in DMEM media and aliquots of 100 μ L were dropped into round areas (prepared using liquid‐repellent slide marker pen) on microscopic glasses, followed by confocal microscopy analysis as described (Garcia‐Faroldi et al. ).…”