Methyl-CpG-binding (SmMBD2/3) and chromobox (SmCBX) proteins are required for neoblast proliferation and oviposition in the parasitic blood fluke Schistosoma mansoni

Abstract

While schistosomiasis remains a significant health problem in low to middle income countries, it also represents a recently recognised threat to more economically-developed regions. Until a vaccine is developed, this neglected infectious disease is primarily controlled by praziquantel, a drug with a currently unknown mechanism of action. By further elucidating how Schistosoma molecular components cooperate to regulate parasite developmental processes, next generation targets will be identified. Here, we continue our studies on schistosome epigenetic participants and characterise the function of a DNA methylation reader, the Schistosoma mansoni methyl-CpG-binding domain protein (SmMBD2/3). Firstly, we demonstrate that SmMBD2/3 contains amino acid features essential for 5-methyl cytosine (5mC) binding and illustrate that adult schistosome nuclear extracts (females > males) contain this activity. We subsequently show that SmMBD2/3 translocates into nuclear compartments of transfected murine NIH-3T3 fibroblasts and recombinant SmMBD2/3 exhibits 5mC binding activity. Secondly, using a yeast-two hybrid (Y2H) screen, we show that SmMBD2/3 interacts with the chromo shadow domain (CSD) of an epigenetic adaptor, S. mansoni chromobox protein (SmCBX). Moreover, fluorescent in situ hybridisation (FISH) mediated co-localisation of Smmbd2/3 and Smcbx to mesenchymal cells as well as somatic- and reproductive- stem cells confirms the Y2H results and demonstrates that these interacting partners are ubiquitously expressed and found within both differentiated as well as proliferating cells. Finally, using RNA interference, we reveal that depletion of Smmbd2/3 or Smcbx in adult females leads to significant reductions (46–58%) in the number of proliferating somatic stem cells (PSCs or neoblasts) as well as in the quantity of in vitro laid eggs. Collectively, these results further expand upon the schistosome components involved in epigenetic processes and suggest that pharmacological inhibition of SmMBD2/3 and/or SmCBX biology could prove useful in the development of future schistosomiasis control strategies.

Schistosoma mansonilysine specific demethylase 1 (SmLSD1) is a druggable target involved in parasite survival, oviposition and stem cell proliferation

“…These proliferation defects are comparable to that observed in Hslsd1-expressing neural stem cells treated with the LSD1 inhibitors pargyline or tranylcypromine [59] and support previous findings confirming that Smlsd1 is also expressed in rapidly dividing cells throughout the parasite [78]. Smmbd2/3 (encoding an epigenetic reader of 5-methylcytosine) and Smcbx (encoding an epigenetic reader of methyl lysine) are also co-expressed in proliferative schistosome cells (h2b+); similar to SmLSD1 inhibition, knockdown of either reader results in reduced neoblast proliferation [79]. When additionally considered alongside neoblast proliferation defects found in adult schistosomes treated with 5-azacytidine (a DNA methyltransferase inhibitor [15,80]), epigenetic processes are rapidly emerging as essential regulators of schistosome stem cell biology.…”

Epigenetic and parasitological parameters are modulated in EBi3-/- mice infected with Schistosoma mansoni

“…SmMBD expression levels were lower in male parasites recovered from knockout animals than in those recovered from wild-type mice. This gene has been linked to normal egg production and number of eggs produced [20], and our EBi3 -/-I mice eliminated smaller quantities of eggs in feces. SmMBD forms part of the NuRD repression complex along with the HDAC8 enzyme [42].…”

“…MBD methyl CpG binding domain is a protein that binds to methylated cytosines, and S. mansoni expressed it at all stages of the parasite's life cycle including both sexes. In the same approach the authors found that this protein has nuclear localization only and relates to oviposition [20]. Previous studies identified 17 USP orthologs in S. mansoni, by homology with 56 USPs present in H. sapiens, it was detected that the expression of these USPs is altered in stages of egg, cercariae, schistosomula and adult worms, reflecting a recycling of proteins used by the parasite during the evolutionary stage transition [21].…”

An Abies procera-derived tetracyclic triterpene containing a steroid-like nucleus core and a lactone side chain attenuates in vitro survival of both Fasciola hepatica and Schistosoma mansoni

“…Adult S. mansoni worms were cultured (as described above) for 72 h in a sub lethal concentration of 700015 (10 μM). After 72 h, a 1 μL aliquot of 10 mM EdU was added to the culture media and incubated for a further 24 h. Worms were subsequently collected and fixed as described previously ( Collins et al, 2013 ; Geyer et al, 2018 ). Anterior regions of both sexes were imaged on a Leica TCS SP5II confocal microscope using a 40× lens.…”