Heat shock-induced cell death




Chen, Miao-Der

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Hyperthermia or heat shock therapy has been utilized in clinical cancer therapy in combination with surgery, chemotherapy, or radiotherapy for the treatment of various types of cancer. However, the precise mechanisms that mediate heat shock-induced apoptotic signaling remains unclear. In previous studies, we have found that none of the known initiator caspases-1, -2, -4, -8, -9, -10, -12, or their caspase-activating complexes, is essential for promoting mitochondrial outer membrane permeabilization (MOMP) or activating caspase-3 following heat shock. Nevertheless, each of the aforementioned events could be significantly inhibited with the pan-caspase inhibitor Z-VAD-FMK. Proapoptotic BCL-2 family members Bax and Bak are essential for inducing MOMP and mediate apoptosis induced by most types of stressful stimuli. Remarkably, however, caspase-3 was still activated in Bax[superscript -/-] /Bak[superscript -/-] DKO MEFs and about 50 % of cells still died following heat shock, even though they failed to undergo MOMP. This suggested that there is another protease, upstream of mitochondria that could activate caspase-3 following heat shock. In subsequent experiments, we discovered that heat shock induced lysosomal membrane permeabilization (LMP) as detected by loss of LysoTracker®-Green and release of cathepsins. Cells lacking the BH3-only protein Bid, which is a direct activator of Bax/Bak, and often cleaved by lysosomal proteases, were not resistant to heat-induced LMP and cell death. However, cells lacking the BH3-only protein Bim were fully resistant to both LMP and cell death, indicating that both events were Bim-dependent. Since heat shock induced the release of cathepsins into the cytosol and some cathepsins are inhibited by Z-VAD-FMK, we speculate that cathepsins might serve as the apical protease. In agreement of this hypothesis, overexpression of a cysteine cathepsin inhibitor suppressed cell death and knockout of cathepsin L completely blocked it. Thus, Bim mediates Bax/Bak-dependent and Bax/Bak-independent cell death following heat shock, and these pathways involved cathepsin L. More recently, we discovered that Bim mutants (BimG154E and BimΔBH3), that could not activate Bax or Bak, nevertheless mediated heat shock-induced LMP and cell death. Bim[superscript -/-] cells and those expressing a Bim mutant (BimG154E/STA) that could no longer associate with the dynein motor complex were fully resistant to heat shock. Thus, Bim’s ability to induce heat shock-induced LMP and cell death did not require its BH3 domain but did require an interaction with LC8 light chain of dynein motor complex. It has been suggested that Bim is sequestered by LC8 in order to prevent unintentional apoptosis. Moreover, a recent study suggests that Bim and LC8 sequester Becin-1, a key regulator of autophagy. However, knockout of essential autophagy genes ATG5 and ATG16L1 did not significantly enhance cell death. Instead, we found the novel discovery that Bim’s association with the dynein motor complex is important for regulating the trafficking of lysosomes. We observed that loss of Bim resulted in a significant decrease in the number and position of lysosomes within the cell, and that reintroduction of Bim (G154E) or Bim (ΔBH3) but not Bim (G154E/STA), resulted in normal lysosome number and distribution. Finally, changes in lysosomal distribution paralleled changes in the secretion of cathepsins into the extracellular space, and both intracellular and extracellular cathepsins appeared to participate in cell death. Thus, in summary, Bim mediates lysosomal cell death through its regulation of lysosome positioning and cathepsin trafficking


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