Metformin regulates the FoxO activity in Echinococcus sp.

LOOS JULIA .A.; DÁVILA VALERIA A.; CUMINO ANDREA C.

Argel

Congreso; 7th Congress of the World Association of Echinococcosis (WAE); 2017

World Association of Echinococcosis

In a wide range of organisms the Forkhead transcription factor FoxO and the SIRT deacetylase constitute a nutrient-sensing pathway involved in regulation of multiple cellular functions. FoxO proteins function mainly as transcriptional activators by binding the consensus core recognition motif TTGTTTAC, and their activity is inhibited by the insulin and IGF-1 signaling pathway. Conversely, in the absence of growth factor signaling or upon cellular stress, FoxOs translocate into the nucleus and activate FoxO-dependent gene expression. Currently, this pathway remains unknown in Echinocococus sp. We have previously shown that Metformin (Met), an anti-hyperglycemic and anti-proliferative drug, exhibits considerable in vitro and in vivo activity against E. granulosus metacestodes. Here, we extended the study and demonstrated that the drug also possess chemopreventive properties against alveolar echinococcosis in mice. As drug administration was shown to induce the Eg-AMPK activation, its anti-echinococcal effects might be a consequence of cellular energy charge depletion in the parasite. Based on this and the fact that only one FoxO transcription factor is present in the genome of Echinococcus sp., the aim of this work is investigate the activation state of FoxO and its relation with the expression of genes encoding sirtuins and key autophagy-related proteins in parasites incubated under both control and energy-stress conditions. Eg-FoxO sequence reveals several post-translationally modifiable residues highly conserved. By in toto immunolocalization assays, we detected the expression and subcellular localization of a phosphorylated (Ser352) and an acetylated (Lys373) form of Eg-FoxO in control and Met-treated protoscoleces. Interestingly, similar expression patterns were observed in both samples. Additionally, by qPCR analysis, we found that Met produced an increase in the transcriptional expression of sirt and/or atg genes in E. granulosus protoscoleces and metacestodes and in E. multilocularis primary cells. In this regards, BLASTn analysis of the upstream sequences in putative promoters of several of these genes showed the conserved binding motif described for FoxO-activated genes. These results suggest a possible role of FoxO in the transcriptional regulation of Echinococcus sp. under energy stress conditions. We also detected expression of Atg8 polypeptide (LC3) with both a diffuse and punctate staining in control and Met-treated E. granulosus protoscoleces and E. multilocularis vesicles. However, western blot analysis demonstrated higher levels of Eg-Atg8-PE (LC3-II) in Met-treated protoscoleces, suggesting a possible induction of autophagy under this condition. Altogether, our data indicate that FoxO, Sirt and autophagy might participate to some extent in the regulation of Met-stimulated energy stress in Echinococcus spp.