Functional and evolutionary characterization of IRE-XBP1 pathway in Echinococcus sp

Abstract

The alterations in protein folding and assembly can induce theendoplasmic reticulum (ER) UPR. IRE1 is the most evolutionarilyconserved ER stress transducer, which upon activation, undergoesdimerization-dependent auto-phosphorylation, and allostericallyinduces its cytosolic endoribonuclease activity. Successively, IREleads to unconventional splicing of the XBP1 mRNA. The translationof this spliced transcription factor results in the transcriptionalinduction of genes expressing chaperones, ERAD components, andautophagy regulators. Previously, we have identified orthologues ofIRE2, XBP1, and ATF6 in the genome of E. granulosus, the causativeagent of human echinococcosis. In this work, we evolutionaryand functionally characterized the IRE/XBP pathway in Echinococcussp. Based on computational assays, we studied the evolutionaryrelationship of Echinococcus-IRE/XBP proteins among metazoansand performed a phylogenetic tree showing their position. Also, weshowed that Echinococcus-IRE has considerable similarity in secondaryand tertiary structures to human IRE (3p23.1A homodimer,35% identity and 0.49 QMEANDisCo). Moreover, we determinedthat the level of total xbp1 transcript increased under ER-stress inducerstreatment and, consequently provokes the enhancement ofgrp78 mRNA expression indicating the occurrence of ER stress inthe parasite. Splicing of the xbp1 mRNA was analyzed to assessIRE1 activation. Through RT-PCR and sequencing, we corroboratedthe presence of unspliced- and spliced- xbp1 in the parasites. Additionally,since XBP-1 mediates the activation of TFEB, a transcriptionfactor that promotes autophagy, we verified that IRE1/XBP1 activationinduces autophagy, with overexpression of Echinococcus atg6,atg8 and tfeb genes. The crosstalk between the IRE1/XBP1 branchand autophagy highlights the importance of both mechanisms inparasite survival. Therefore, targeting the UPR-induced autophagyresponse may lead to novel therapeutic approaches.