Whole worm microRNA silencing in cestodes

GRECCO, ANDRÉS; MACCHIAROLI, NATALIA; CUCHER, MARCELA; ROSENZVIT, MARA

Nueva York

Congreso; Regulatory and Non-coding RNAs Meeting 2022; 2022

Cold Spring Harbor Laboratory

Mesocestoides vogae (syn.M.corti) is a platyhelminth parasite validated as a laboratory model of the Cestoda class. Cestodes include species such as Echinococcus granulosus and E. multilocularis that cause Tropical Neglected Diseases (NTDs) such as echinococcosis. Although genomic information is available for several of these parasites, functional postgenomic strategies are very limited so far. microRNAs (miRNAs) are small non-coding RNAs with a main role in negative regulation of gene expression as they recognize target sites in the mRNA 3’UTRs. miR-71 is a highly expressed miRNA in cestodes, it is absent in vertebrates and targets essential parasite genes. Thus, miR-71 represents a potential selective drug target to treat cestodiases. In a previous work from our group, in vitro silencing of miR-71 in E. multilocularis cell cultures was successfully performed. In this work we aim to knockdown miR-71 in whole larvae (tetrathyridia) from M. vogae. To this end, a strategy based on antimiRs was selected. AntimiRs are single-stranded oligonucleotides complementary to the miRNA, which block miRNA function. First, transfections with increasing concentrations of a fluorescent scrambled oligonucleotide were performed on cultured tetrathyridia and analyzed through confocal microscopy to determine the best uptake conditions. After that, antimiR transfections were performed in 3 biological replicates. Administration of antimiR-71 resulted in miR-71 knockdown of 65%measured by RT-qPCR. Strobilization of "miR-71 silenced" and control tetrathyridia was induced, strobilization rates for the silenced group were higher. These results suggest that miR-71 might be involved in the negative regulation of strobilization associated genes. Experimental validation of miR-71 target genes in silico predicted and analysis of miRNA silencing effects on parasite development are underway. This results provide a strategy to study miRNA function in cestodes and to analyze miRNA potential as drug targets against NTDs.