Vacuolar (H⁺)‐ATPase subunit c is essential for the survival and systemic RNA interference response in Locusta migratoria

Abstract

Background: Vacuolar (H + )-ATPase (V-ATPase) is a multi-subunit enzyme that hydrolyzes adenosine triphosphate (ATP) to transport protons across a cellular membrane, and it plays an important role in numerous biological processes, including in growth, development and immune responses. The c subunit of V-ATPase is a highly conserved subunit of the rotatory proteolipid ring that is required for binding and transporting protons. To date, there are only a few published reports on V-ATPase-c functions in insects.Results: We identified and characterized the V-ATPase-c gene in Locusta migratoria, one of the most destructive agricultural insect pests in the world. LmV-ATPase-c was predominately expressed in Malpighian tubules of nymphs, followed by the hindgut and ovary, while the other tissues showed relatively low expression levels. Silencing of LmV-ATPase-c caused severe molting defects in nymphs and a high mortality rate of > 90%. Histological staining and microscopic examination of sections from the abdominal cuticle revealed the absence of newly formed cuticle in nymphs that were injected with dsLmV-ATPase-c. In addition, silencing of LmV-ATPase-c transcript levels significantly impaired RNA interference (RNAi) efficiency of a reporter gene. By quantifying double-stranded RNA (dsRNA) amounts by quantitative polymerase chain reaction (PCR), we found that RNAi against LmV-ATPase-c provoked a dramatic accumulation of dsRNA in the endosomes of epidermal and midgut cells of Locusta migratoria. Conclusion:Our results indicate that LmV-ATPase-c is indispensable for the formation of new cuticle during the molting process and has pivotal functions in dsRNA escape from endosomes. LmV-ATPase-c might be a valuable target for developing new strategies for insect pest management.

Novel Environmentally Friendly RNAi Biopesticides: Targeting V-ATPase in Holotrichia parallela Larvae Using Layered Double Hydroxide Nanocomplexes

“…V-ATPase , widely distributed in the plasma membrane of epithelial cells such as Malpighian tubules, midgut, and salivary glands in insects, plays a vital role in intestinal alkalization, nutrient absorption, and overall insect growth and development. It represents a potential RNAi candidate for biological control. , This study focused on silencing the HpV-ATPase gene, resulting in an 86.67% larval mortality rate, underscoring its significance in the growth and development of H. parallela larvae.…”

“…It has currently been studied and applied in various insects, serving as a potential RNAi candidate for biological control. − Silencing the LmV-ATPase-a gene in L. migratoria resulted in a 98.6% insect mortality rate, while V-ATPase-c caused severe molting defects in nymphs with a mortality rate >90% . Using RNAi to interfere with Eriocheir sinensis V-ATPase subunit B ( EsVATB ) revealed a high mortality rate (74%), a low molting rate (26%), and a failure to form a new epicuticle in the premolt stage of crabs …”

Bacteria‐mediated RNAi for managing fall webworm, Hyphantria cunea: screening target genes and analyzing lethal effect

“…Regardless of the cellular uptake of dsRNA or the systemic spreading of RNAi, dsRNAs are transferred through vesicles 40 . vATPase is responsible for the endocytosis‐mediated uptake of dsRNA in multiple insects 66,67 . Whether these genes are involved in dsRNA transport in H. cunea requires further investigation.…”

“…40 vATPase is responsible for the endocytosis-mediated uptake of dsRNA in multiple insects. 66,67 Whether these genes are involved in dsRNA transport in H. cunea requires further investigation.…”

Heterologous expression of circular RNAs in Escherichia coli for analyzing the ligation process of chloroplastic viroids and producing double-stranded RNAs with insecticidal activity

“…Blocking the interaction of late endosomes with lysosomes enhances si-and miRNA-mediated silencing in D. melanogaster, while blocking late endosome formation limits silencing (Lee et al, 2009), thus restricting dsRNA escape between late endosome formation and lysosomal fusion . The activity of the vacuolar H + -ATPase has been related to dsRNA cell entry in several species Xiao et al, 2015;Shi et al, 2022); however, the mechanism mediating the endosomal escape is not exactly known.…”