Ceramidases (CDases) are vital enzymes involved in the biosynthesis of sphingolipids, which are essential components of eukaryotic membranes. The function of these enzymes in insects, however, is poorly understood. We identified a neutral ceramidase (NlnCDase) from the brown planthopper, Nilaparvata lugens, one of the most destructive hemipteran pests of rice. The C12-ceramide was the most preferred substrate for the NlnCDase enzyme. The activity of the NlnCDase enzyme was highest in the neutral-pH range (pH 6.0). It was inhibited by EGTA, Cs+ and Fe2+, while stimulated by EDTA and Ca2+. Moreover, the NlnCDase has higher transcript level and activity in adults than in eggs and nymphs, and in the reproductive organs (ovaries and spermaries) than in other tissues (i.e. heads, thorax, legs, midguts), which suggested that the NlnCDase might be elevated to mediate developmental process. In addition, transcripts and activity of the NlnCDase were up-regulated under abiotic stresses including starvation, abnormal temperature, and insecticides, and biotic stress of resistant rice varieties. Knocking down NlnCDase by RNA interference increased female survival under starvation and temperature stresses, suggesting that NlnCDase might be involved in the stress response in N. lugens.

The neutral sphingomyelinase (nSMase) 1 homologue gene LsSMase was cloned from Laodelphax striatellus, a direct sap-sucker and virus vector of gramineous plants, and expressed via a Bac to Bac baculovirus expression system. The LsSMase-enhanced green fluorescent protein fusion protein was located in the endoplasmic reticulum in a similar manner to mammalian nSMase 1. The biochemical properties of LsSMase were determined in detail. The optimal pH and temperature for recombinant LsSMase were 8 and 37 °C, respectively. LsSMase was an Mg or Mn dependent enzyme, but different concentration of each were needed. The activity of LsSMase was significantly stimulated by Ethylene glycol bis(2-aminoethyl ether)tetraacetic acid (EGTA), whereas it was inhibited by ethylenediaminetetraacetic acid. Millimolar concentrations of Zn completely inhibited LsSMase. The reducing agents dithiothreitol and β-mercaptoethanol varied in their effects on activity. Phospholipids were not found to stimulate LsSMase.