Knockdown of Vacuolar ATPase Subunit G Gene Affects Larval Survival and Impaired Pupation and Adult Emergence in Henosepilachna vigintioctopunctata

SIMPLE SUMMARY: Vacuolar ATPase (vATPase), a proton pump driven by ATP hydrolysis, acts as a membrane energizer to motivate the movement of ions and nutrients across the cellular membrane in insect guts and Malpighian tubules, among others. The vATPase holoenzyme contains 16 subunits. Out of these subunits, mammalian G subunit includes three isoforms (G1-G3) which are encoded by three distinctive genes. The physiological role of a specific G isoform can be compensated by others. Thus, current experimental evidence on the in vivo function of G is rather limited among eight V(1) subunits. In the present paper, particular attention was paid to an insect model, Henosepilachna vigintioctopunctata ladybird, a serious defoliator of Solanaceae and Cucurbitaceae plants in many Asian countries. Given that the beetle is sensitive to RNA interference (RNAi), HvvATPaseG gene was knocked down by ingestion of its corresponding dsRNA at the fourth-instar larval stage. Silence of HvvATPaseG affected larval growth and survival, impaired pupation and adult emergence. Our results provide a basis for further functional research on the vATPase G subunit in insects and suggest new ideas for the management of H. vigintioctopunctata. ABSTRACT: The vATPase holoenzyme consists of two functional subcomplexes, the cytoplasmic (peripheral) V(1) and the membrane-embedded V(0). Both V(1) and V(0) sectors contain eight subunits, with stoichiometry of A(3)B(3)CDE(3)FG(3)H in V(1) and ac(8)c’c”def(Voa1p) in V(0) respectively. However, the function of G subunit has not been characterized in any non-Drosophilid insect species. In the present paper, we uncovered that HvvATPaseG was actively transcribed from embryo to adult in a Coleopteran pest Henosepilachna vigintioctopunctata. Its mRNA levels peaked in larval hindgut and Malpighian tubules. RNA interference (RNAi)-mediated knockdown of HvvATPaseG significantly reduced larval feeding, affected chitin biosynthesis, destroyed midgut integrity, damaged midgut peritrophic membrane, and retarded larval growth. The function of Malpighian tubules was damaged, the contents of glucose, trehalose, lipid, total soluble amino acids and protein were lowered and the fat bodies were lessened in the HvvATPaseG RNAi larvae, compared with those in the PBS- and dsegfp-fed beetles. In contrast, the amount of glycogen was dramatically increased in the HvvATPaseG depletion ladybirds. As a result, the development was arrested, pupation was inhibited and adult emergence was impaired in the HvvATPaseG hypomorphs. Our results demonstrated that G subunit plays a critical role during larval development in H. vigintioctopunctata.