The LdCDF4 confers Cu tolerance of Lymantria dispar larvae: A novel heavy metal transporter in insects

Abstract

Even though copper (Cu) is a necessary trace element, it can cause growth toxicity in insects. This study investigates the mechanism of Cu tolerance in Lymantria dispar larvae in relation to the cation diffusion facilitator family (CDF). The results showed that larval mortality of L. dispar exhibited a dose-dependent response to Cu exposure, with a survival rate of 43 % even at high Cu concentrations (500 mg/kg). Of the seven CDF family genes examined, only LdCDF4 showed significant upregulation in L. dispar larvae following Cu treatment. Subcellular localization experiments indicated that the LdCDF4 protein localized to the cell membrane. At the individual level, compared with the L. dispar larvae treated with Cu alone, silencing of LdCDF4 under Cu stress led to reduced body weight, prolonged developmental duration, upregulation of apoptosis-related genes, and further disruption of genes in the mitochondrial apoptosis pathway. At the cellular level, LdCDF4 overexpression mitigated Cu-induced damage in Sf9 cells by enhancing cell viability, decreasing apoptosis, lowering Ca²⁺ levels, reducing reactive oxygen species (ROS) production, minimizing mitochondrial permeability transition pore (MPTP) opening, and maintaining mitochondrial membrane potential. Collectively, L. dispar larvae exhibited strong Cu tolerance, with LdCDF4 playing a key role by alleviating the ROS/Ca²⁺-MPTP opening-mitochondrial membrane potential-apoptosis cascade.