Functional analysis of male competence factors potentially involved in the male-female interaction of Schistosoma mansoni

Abstract text

Introduction

Schistosoma mansoni is a parasite that causes schistosomiasis, a neglected tropical disease. Schistosomes are unique in their reproductive biology, because the female depends on a constant pairing contact with a male partner to achieve sexual maturation. Although there are reports on functional description of GPCRs and their role in metabolic pathways in vertebrates, mammals and plants, roles of GPCRs in invertebrates are not well studied.

Objective

Our research focuses on the functional characterization of genes coding for male competence factors that may regulate the sexual maturation of the female after pairing in S. mansoni. To this end, we identified pairing-dependently expressed GPCRs and neuropeptide genes (npps) that are upregulated in males and downregulated in females after pairing. To unravel GPCR-npp interaction, we established MALAR-Y2H system, which provided first evidence for matching GPCR-npp couples. Certain GPCR-npp interactions have already been identified in previous studies of our group.

Materials and Methods

Here, I studied the function of GPCR 21, which was downregulated by RNA interference (RNAi) experiments in vitro. The experiment was performed in pairing-experienced (bsF) and inexperienced females (ssF) followed by repairing. RNAi effects on pairing stability, egg production, motility, and attachment behavior were monitored as physiological parameters. In addition, the localization of the GPCR 21 transcript was investigated by whole mount in situ hybridization (WISH). Finally, the morphological changes with respect to knock down were analyzed using microscopy.

Results and Conclusion

A male and ssF-dominated transcript profile of GPCR 21 was identified and reconfirmed earlier by our group. WISH localized transcripts were seen in the anterior region and the gut area of adult worms. The RNAi-mediated knockdown efficiencies for bsF and ssF were 39% and 64%, respectively. An increased percentage in attachment was observed after repairing. In both the bsF and ssF groups, an increased percentage in attachment was observed after repairing as well as a reduction in motility over time of the experiment. I observed an additional negative effect on egg production in the bsF group. I plan to do further investigation on identifying the npp interaction partner of GPCR 21 and will perform FRET analysis for its confirmation. The efficiency of RNAi experiments will be improved by performing a double knockdown.