4.7 Article

Rapid cold-hardening protects Drosophila melanogaster from cold-induced apoptosis

Journal

APOPTOSIS
Volume 12, Issue 7, Pages 1183-1193

Publisher

SPRINGER
DOI: 10.1007/s10495-006-0048-2

Keywords

Drosophila melanogaster; apoptosis; TUNEL assay; rapid cold-hardening; cold shock

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The rapid cold-hardening (RCH) response increases the cold tolerance of insects by protecting against non-freezing, cold-shock injury. Apoptosis, or programmed cell death, plays important roles in development and the elimination of sub-lethally damaged cells. Our objectives were to determine whether apoptosis plays a role in cold-shock injury and, if so, whether the RCH response protects against cold-induced apoptosis in Drosophila melanogaster. The present study confirmed that RCH increased the cold tolerance of the adults at the organismal level. No flies in the cold-shocked group survived direct exposure to similar to 7 degrees C for 2 h, whereas significantly more flies in the RCH group survived exposure to -7 degrees C for 2 h after a 2-h exposure to 5 degrees C. We used a TUNEL assay to detect and quantify apoptotic cell death in five groups of flies including control, cold-shocked, RCH, heat-shocked (37.5 degrees C, 30 min), and frozen (-20 degrees C, 24 h) and found that apoptosis was induced by cold shock, heat shock, and freezing. The RCH treatment significantly improved cell viability by 38% compared to the cold-shocked group. Cold shock-induced DNA fragmentation shown by electrophoresis provided further evidence for apoptosis. SDS-PAGE analysis revealed an RCH-specific protein band with molecular mass of similar to 150 kDa. Western-blotting revealed three proteins that play key roles in the apoptotic pathway: caspase-9-like (apoptotic initiator), caspase-3-like (apoptotic executioner) and Bcl-2 (anti-apoptotic protein). Consequently, the results of this study support the hypothesis that the RCH response protects against cold-shock-induced apoptosis.

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