Serine/threonine protein phosphatase 1 (PP1) controls growth and reproduction in Schistosoma japonicum.

Schistosomiasis is a human parasitic disease caused by flatworms of the genus Schistosoma. Adult female schistosomes produce numerous eggs that are responsible for the pathogenesis and transmission of the disease, and the maturation of female gonads depends on the permanent pairing of females and males. Signaling protein kinases have been proven to control female gonad differentiation after pairing; however, little is known about the roles of protein phosphatases in the developmental and reproductive biology of schistosomes. Here we explored 3 genes encoding catalytic subunits of serine/threonine protein phosphatase 1 (PP1c) that were structurally and evolutionarily conserved in Schistosoma japonicum. In situ hybridization showed transcripts of 3 Sj-pp1c genes mainly localized in the reproductive organs and tissues. Triple knockdown of Sj-pp1c genes by RNA interference caused stunted growth and decreased pairing stability of worm pairs, as well as a remarkable reduction in cell proliferation activity and defects in reproductive maturation and fecundity. Transcriptomic analysis post-RNA interference suggested that Sj-pp1c genes are involved in controlling worm development and maturation mainly by regulating cell proliferation, eggshell synthesis, nutritional metabolism, cytoskeleton organization, and neural process. Our study provides the first insight into the fundamental contribution of Sj-PP1c to molecular mechanisms underlying the reproductive biology of schistosomes.-Zhao, L., Lu, Z., He, X., Mughal, M. N., Fang, R., Zhou, Y., Zhao, J., Gasser, R. B., Grevelding, C. G., Ye, Q., Hu, M. Serine/threonine protein phosphatase 1 (PP1) controls growth and reproduction in Schistosoma japonicum.

The ABL kinase inhibitor imatinib causes phenotypic changes and lethality in adult Schistosoma japonicum.

Schistosomiasis caused by different species of schistosome parasites is one of the most debilitating helminthic diseases of humans worldwide. For decades, chemotherapy is the main method of controlling schistosomiasis. However, the fear of drug resistance has motivated the search for alternatives. It has been demonstrated that the ABL kinase inhibitor imatinib affected the development and survival of Schistosoma mansoni in vitro; however, there is still lack of information on whether imatinib also affects other schistosome species such as Schistosoma japonicum. In the present study, the anti-schistosomal potency of imatinib on adult S. japonicum was investigated in vitro, and the results showed that imatinib had a significant impact on various physiological processes of S. japonicum adult worms. Besides its negative effects on worm motility, pairing stability, and gonad development, imatinib caused pathological changes in the gastrodermis as well as the death of the parasite. Our findings suggest that imatinib is an intriguing candidate for further development as an option to fight S. japonicum.

The anticancer drug imatinib induces autophagy in Schistosoma mansoni.

Schistosomiasis, caused by schistosome parasites, is a neglected tropical disease affecting humans and animals. There is no vaccine available yet, and fear of upcoming resistance against the only widely used drug, praziquantel, is omnipresent. Previously, we showed that imatinib (Gleevec), an anticancer drug, affected schistosome physiology and caused the death of adult Schistosoma mansoni in vitro. Here, we present the first known evidence that one effect of imatinib is the induction of autophagy in S. mansoni. Furthermore, worms co-treated with imatinib and bafilomycin A1, a late-phase autophagy inhibitor, reversed imatinib-induced autophagy and its antischistosomal effects as revealed by phenotypic and molecular analyses.

First insights into the autophagy machinery of adult Schistosoma mansoni.

Schistosomiasis is a disease of global importance caused by parasitic flatworms, schistosomes, which cause pathogenicity through eggs laid by the female worm inside the host's blood vessels. Maintenance of cellular homeostasis is crucial for parasites, as for other organisms, and is quite likely important for schistosome reproduction and vitality. We hypothesize a role for autophagy in these processes, an evolutionarily conserved and essential cellular degradation pathway. Here, for the first known time, we shed light on the autophagy machinery and its involvement in pairing-dependent processes, vitality and reproduction of Schistosoma mansoni. We identified autophagy genes by in silico analyses and determined the influence of in vitro culture on the transcriptional expression in male and female worms using quantitative real-time PCR. Among the identified autophagy genes were Beclin, Ambra1, Vps34, DRAM, DAP1, and LC3B, of which some showed a sex-dependent expression. Specifically, the death-associated protein DAP1 was significantly more highly expressed in females compared with males, while for the damage-regulated autophagy modulator DRAM it was the opposite. Furthermore, in-vitro culture significantly changed the transcript expression level of DAP1 in female worms. Next, worms were treated with an autophagy inducer (rapamycin) or inhibitors (bafilomycin A1, wortmannin and spautin-1) to evaluate effects on autophagy protein expression, worm vitality, and reproduction. The conversion of the key autophagy protein LC3B, a marker for autophagic activity, was increased by rapamycin and blocked by bafilomycin. All inhibitors affected worm fitness, egg production, and negatively affected the morphology of gonads and intestine. In summary, autophagy genes in S. mansoni show an interesting sex-dependent expression pattern and manipulation of autophagy in S. mansoni by inhibitors induced detrimental effects, which encourages subsequent studies to identify antischistosomal targets within the autophagy machinery.

First Evidence of Function for Schistosoma japonicumriok-1 and RIOK-1.

Protein kinases are known as key molecules that regulate many biological processes in animals. The right open reading frame protein kinase (riok) genes are known to be essential regulators in model organisms such as the free-living nematode Caenorhabditis elegans. However, very little is known about their function in parasitic trematodes (flukes). In the present study, we characterized the riok-1 gene (Sj-riok-1) and the inferred protein (Sj-RIOK-1) in the parasitic blood fluke, Schistosoma japonicum. We gained a first insight into function of this gene/protein through double-stranded RNA interference (RNAi) and chemical inhibition. RNAi significantly reduced Sj-riok-1 transcription in both female and male worms compared with untreated control worms, and subtle morphological alterations were detected in the ovaries of female worms. Chemical knockdown of Sj-RIOK-1 with toyocamycin (a specific RIOK-1 inhibitor/probe) caused a substantial reduction in worm viability and a major accumulation of mature oocytes in the seminal receptacle (female worms), and of spermatozoa in the sperm vesicle (male worms). These phenotypic alterations indicate that the function of Sj-riok-1 is linked to developmental and/or reproductive processes in S. japonicum.

The RIO protein kinase-encoding gene Sj-riok-2 is involved in key reproductive processes in Schistosoma japonicum.

BACKGROUND: Schistosomiasis is one of the most prevalent parasitic diseases worldwide and is caused by parasitic trematodes of the genus Schistosoma. The pathogenesis of schistosomiasis is caused by eggs whose production is the consequence of the pairing of schistosomes and the subsequent sexual maturation of the female. Previous studies have demonstrated that protein kinases are involved in processes leading to the male-induced differentiation of the female gonads, ovary and vitellarium. Right open reading frame protein kinase 2 (RIOK-2) is a member of the atypical kinase family and shown in other organisms to be responsible for ribosomal RNA biogenesis and cell-cycle progression, as well as involves in nematode development. However, nothing is known about its functions in any trematode including schistosome. METHODS: We isolated and characterized the riok-2 gene from S. japonicum, and detected the transcriptional profiles of Sj-riok-2 by using real-time PCR and in situ hybridization. RNAi-mediated knockdown of Sj-riok-2 was performed, mitotic activities were detected by EdU incorporation assay and morphological changes on organs were observed by confocal laser scanning microscope (CLSM). RESULTS: In silico analyses of the amino acid sequence of Sj-RIOK-2 revealed typical features of this class of kinases including a winged helix (wHTH) domain and a RIO kinase domain. Sj-riok-2 is transcribed in different developmental stages of S. japonicum, with a higher abundance in adult females and eggs. Localization studies showed that Sj-riok-2 was mainly transcribed in female reproductive organs. Experiments with adult schistosomes in vitro demonstrated that the transcriptional level of Sj-riok-2 was affected by pairing. Knocking down Sj-riok-2 by RNAi reduced cell proliferation in the vitellarium and caused the increased amount of mature oocytes in ovary and an accumulation of eggs within the uterus. CONCLUSIONS: Sj-riok-2 is involved in the reproductive development and maturation of female S. japonicum. Our findings provide first evidence for a pairing-dependent role of Sj-riok-2 in the reproductive development and maturation of female S. japonicum. Thus this study contributes to the understanding of molecular processes controlling reproduction in schistosomes.