Rapid cold-hardening blocks cold-induced apoptosis by inhibiting the activation of pro-caspases in the flesh fly Sarcophaga crassipalpis

Apoptosis plays important roles in the selective elimination of sub-lethally damaged cells due to various environmental stresses. The rapid cold-hardening (RCH) response protects insects from the otherwise lethal consequences of injury due to cold-shock. We recently demonstrated that cold shock induces apoptotic cell death in insects and that RCH functions to specifically block cold-shock-induced apoptosis. In the present study we used isolated fat body, midgut, muscle, and Malpighian tubules from adult flesh flies Sarcophaga crassipalpis to test the following hypotheses: (1) cold-induced apoptosis varies among different tissues and (2) RCH blocks the apoptotic pathway by preventing the activation of pro-caspases. Cold-shock induced substantial amounts of apoptotic cell death that matched with tissue damage as determined using vital dyes. RCH treatment significantly reduced apoptotic cell death in all tested tissues. Caspase-3 (executioner) activity was 2-3 times higher in the cold- and heat-shocked groups than in control and RCH groups. Likewise, the activity of caspase-9 (initiator) showed a similar trend as for caspase-3 in all tissues but midgut. In addition, cold-shock and heat-shock treatments also increased caspase-2 activity 2-3 folds in both soluble and membrane fractions of fat body and muscle extracts compared to controls.