MOORE, C. W.*; YI, S-X.; LEE, JR., R. E.; Miami University; Miami University; Miami University: Does the rapid cold-hardening response protect Drosophila from cold-induced apoptosis?

The rapid cold-hardening (RCH) response increases the cold tolerance of insects by protecting against non-freezing, cold-shock injury. Although the RCH response is well documented in a variety of arthropods, little is known about the underlying mechanism that protects against cold-shock injury. Apoptosis, or programmed cell death, plays important roles in development and in 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 prevents this cold-induced apoptosis. RCH increased the cold tolerance of adult Drosophila melanogaster. No flies in the cold-shocked group survived direct exposure to -7°C for 2 h, whereas significantly more flies in the RCH group survived exposure to 5°C for 2 h prior to a 2 h exposure to -7°C. Since apoptosis is characterized by nonrandom DNA fragmentation into ~180-200 base pairs, we extracted DNA from three groups of adult flies: the control (untreated at 23°C), the cold-shocked, and the RCH. DNA gel electrophoresis showed that both control and RCH flies primarily exhibited banding in the 400-600 bp region. In contrast, DNA from the cold-shocked group only displayed heavy banding in the 150-200 bp region, a pattern that is indicative of apoptosis. To determine whether certain proteins were associated with the RCH protection against apoptosis, we extracted protein samples from each group. Our preliminary electrophoretic data indicates the presence of a novel protein band in the RCH group. Overall, our initial results suggest that the RCH response blocks cold-induced apoptosis.