Identifying and validating the presence of Guanine-Quadruplexes (G4) within the blood fluke parasite Schistosoma mansoni

Abstract

Schistosomiasis is a neglected tropical disease that currently affects over 250 million individuals worldwide. In the absence of an immunoprophylactic vaccine and the recognition that mono-chemotherapeutic control of schistosomiasis by praziquantel has limitations, new strategies for managing disease burden are urgently needed. A better understanding of schistosome biology could identify previously undocumented areas suitable for the development of novel interventions. Here, for the first time, we detail the presence of G-quadruplexes (G4) and putative quadruplex forming sequences (PQS) within the Schistosoma mansoni genome. We find that G4 are present in both intragenic and intergenic regions of the seven autosomes as well as the sex-defining allosome pair. Amongst intragenic regions, G4 are particularly enriched in 3´ UTR regions. Gene Ontology (GO) term analysis evidenced significant G4 enrichment in the wnt signalling pathway (p<0.05) and PQS oligonucleotides synthetically derived from wnt-related genes resolve into parallel and anti-parallel G4 motifs as elucidated by circular dichroism (CD) spectroscopy. Finally, utilising a single chain anti-G4 antibody called BG4, we confirm the in situ presence of G4 within both adult female and male worm nuclei. These results collectively suggest that G4-targeted compounds could be tested as novel anthelmintic agents and highlights the possibility that G4-stabilizing molecules could be progressed as candidates for the treatment of schistosomiasis.

G-quadruplexes in helminth parasites

“…We first verified that the S. mansoni G4 sequences recently described in ( 29 ) formed G4s, using the same techniques as the ones described for A. lumbricoides . As shown in Supplementary Figure S5 , this is indeed the case for the five positive sequences described in this article.…”

“…The FRET-MC method measures the ability of a sequence to compete for binding to a well-known G4 ligand, PhenDC3. This compound is highly selective for G4s: efficient competitors are able to act as decoys for this G4 ligand, leading to a strong decrease in ΔT m of a fluorescent G4-forming oligonucleotide ( 29 ); when added in large excess, these specific competitors can lead to negligible Δ T m values. As can be seen in this panel, 19 out of 20 sequences considered here acted as efficient competitors (AL1 was found to be less efficient), arguing for G4 formation for most motifs.…”

“…We next investigated whether G4 ligands (i.e. small compounds which selectively recognize this unusual nucleic acid structure) recently synthesized (Figure 6A ) (35–36) would bind to the S. mansoni G4s described in ( 29 ). Six compounds were chosen, with variable levels of stabilization of G4 structures.…”

“…Comparing the genomes of parasites from these two phyla with different infectivity (parasitic and non-parasitic worms) may reveal details of processes driving pathogenicity and new possible drug targets. Finally, we showed that the recently-identified G4 motifs found in S. mansoni ( 29 ) may be targeted by G4 ligands, leading to anti-parasitic effects.…”

“…A study conducted on A. lumbricoides shows that the fluorescent G4 ligand Q1 binds selectively to the antiparallel telomeric G4 ( 28 ). Very recently, G4 motifs were identified in the genome of S. mansoni and the authors confirmed the presence of G4 in adult worms, by means of the BG4 G-quadruplex-specific antibody ( 29 ).…”