Downregulated RPS-30 in Angiostrongylus cantonensis L5 plays a defensive role against damage due to oxidative stress

Abstract

Background Eosinophilic meningitis, caused by fifth-stage larvae of the nematode (roundworm) Angiostrongylus cantonensis, is mainly attributed to the contribution of eosinophils to tissue inflammatory responses in helminthic infections. Eosinophils are associated with the killing of helminths via peroxidative oxidation and hydrogen peroxide generated by the dismutation of superoxide produced during respiratory bursts. In contrast, when residing in the host with high level of eosinophils, helminthic worms have evolved to attenuate eosinophil-mediated tissue inflammatory responses for their survival in the hosts. In a previous study we demonstrated that the expression of the A. cantonensis RPS 30 gene (Acan-rps-30) was significantly downregulated in A. cantonensis L5 roundworms residing in cerebrospinal fluid with a high level of eosinophils. Acan-RPS-30 is a protein homologous to the human Fau protein that plays a pro-apoptotic regulatory role and may function in protecting worms from oxidative stress. Methods The isolation and structural characterization of Acan-RPS-30 were performed using rapid amplification of cDNA ends (RACE), genome walking and bioinformatics. Quantitative real-time-PCR and microinjection were used to detect the expression patterns of Acan-rps-30. Feeding RNA interference (RNAi) was used to knockdown the apoptosis gene ced-3. Microinjection was performed to construct transgenic worms. An oxidative stress assay was used to determine the functions of Acan-RPS-30. Results Our results showed that Acan-RPS-30 consisted of 130 amino acids. It was grouped into clade V with C. elegans in the phylogenetic analysis. It was expressed ubiquitously in worms and was downregulated in both L5 larvae and adult A. cantonensis. Worms expressing pCe-rps30::Acan-rps-30::rfp, with the refractile “button-like” apoptotic corpses, were susceptible to oxidative stress. Apoptosis genes ced-3 and ced-4 were both upregulated in the transgenic worms. The phenotype susceptible to oxidative stress could be converted with a ced-3 defective mutation and RNAi. rps-30−/− mutant worms were resistant to oxidative stress, with ced-3 and ced-4 both downregulated. The oxidative stress-resistant phenotype could be rescued and inhibited by through the expression of pCe-rps30::Acan-rps-30::rfp in rps-3−/− mutant worms. Conclusion In C. elegans worms, downregulated RPS-30 plays a defensive role against damage due to oxidative stress, facilitating worm survival by regulating downregulated ced-3. This observation may indicate the mechanism by which A. cantonensis L5 worms, with downregulated Acan-RPS-30, survive in the central nervous system of humans from the immune response of eosinophils. Graphic abstract

Up-Regulated Galectin-1 in Angiostrongylus Cantonensis L5 Reduces Body Fat and Increases Oxidative Stress Tolerance

“…Therefore, Eosinophils are the robust producers of extracellular superoxide. In contrast, helminthic worms have evolved to attenuate the oxidative stress damage from eosinophils for their survival in hosts, such as inducing apoptosis of eosinophils [52,53] and increasing oxidative stress resistance of worm itself [17]. We have demonstrated that Acan-Gal-1, up-regulated in A. cantonensis L5, that resides in human with the senior immune system, could induced the apoptosis of macrophages extracellular, when it was secreted by cells via an unconventional mechanism [21,22].…”

“…Primers used for PCR analysis are listed in Additional le 1: Table S1. RNAi plates and media were prepared according to [17]. Gravid adult ced-3 mutant worms and fat-6;fat-7 double-mutant worms were allowed to lay eggs overnight on the RNAi plates and adult worms were picked off.…”

“…As we have demonstrated that down-regulated Acan-RPS-30 in A. cantonensis L5 could resist oxidative stress damage through inhibiting worm apoptosis [17], and oxidative stress is thought to be one of the major factors that promote apoptosis [45], we next determined whether Acan-Gal-1 increased oxidative stress tolerance via inhibiting worm apoptosis. Then, the expression levels of apoptosis genes were detected in C. elegans.…”

“…In our previous study, proteomic analysis of A. cantonensis L5 and A. cantonensis iL3, using twodimensional difference gel electrophoresis (2D-DIGE), showed that the expression level of A. cantonensis RPS-30 (Acan-RPS-30) was lower in L5 than that in iL3, and that of A. cantonensis Galectin-1 (Acan-Gal-1) was higher [8]. Recently, we have demonstrated that down-regulated Acan-RPS-30 in A. cantonensis L5 played a defensive role against damage due to oxidative stress for worm survival through inhibiting apoptosis by regulating ced-3 down-regulated [17].…”

Upregulated galectin-1 in Angiostrongylus cantonensis L5 reduces body fat and increases oxidative stress tolerance

“…In our previous study, proteomic analysis of A. cantonensis L5 and A. cantonensis iL3, using two-dimensional difference gel electrophoresis (2D-DIGE), showed that the expression level of A. cantonensis RPS-30 ( Acan -RPS-30) was lower in L5 than that in iL3 and that of A. cantonensis Galectin-1 ( Acan -Gal-1) was higher [ 8 ]. Recently, we have demonstrated that downregulated Acan- RPS-30 in A. cantonensis L5 plays a defensive role in worm survival against damage due to oxidative stress by inhibiting apoptosis by regulating ced-3 downregulation [ 17 ].…”