RNA interference of glycerol biosynthesis suppresses a rapid cold hardiness of the beet armyworm, Spodoptera exigua

A freeze-susceptible species of the beet armyworm, Spodoptera exigua, overwinters in temperate zones without diapauses. The rapid cold hardening (RCH) and supercooling may play a crucial role in the overwintering. This study determined a cryoprotectant as a rapid cold hardening factor of S. exigua. A pre-exposure of S. exigua larvae to 4°C significantly increased survival at -10°C in all developmental stages from egg to adult. The RCH was dependent on the duration of the pre-exposure period. The RCH also significantly enhanced the supercooling capacity. Cryoprotectant analysis using an HPLC showed that the pre-exposure treatment allowed the larvae to accumulate glycerol in the hemolymph. Two genes of glycerol-3-phosphate dehydrogenase (GPDH) and glycerol kinase (GK) were chosen to be associated with glycerol biosynthesis and cloned in S. exigua larvae. Both GPDH and GK were expressed in all developmental stages of S. exigua. Either RNA interference (RNAi) of GPDH or GK significantly inhibited the glycerol accumulation in the hemolymph of S. exigua. Larvae treated with RNAi of GPDH or GK gene expression exhibited a significant decrease in the RCH ability. The glycerol accumulation in response to 4°C appeared to be under the control by humoral signal because a ligation experiment prevented the glycerol accumulation in the other halve. This study indicated that glycerol is a RCH factor of S. exigua and induced to be synthesized in response to low temperature via humoral mediation.