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

DIAZ, MALENA; LEDO, CAMILA; NICOLAO, MARÍA CELESTE; FORNASARI, M. SILVINA; CUMINO, ANDREA C.

Mar del Plata

Congreso; LXVII Reunión Anual Sociedad Argentina de Investigación Clínica, Reunión Anual de Sociedades de Biociencias (SAIC-SAI-FAIC-SAFIS); 2022

The alterations in protein folding and assembly can induce the endoplasmic reticulum (ER) UPR. IRE1 is the most evolutionarily conserved ER stress transducer, which upon activation, undergoes dimerization-dependent auto-phosphorylation, and allosterically induces its cytosolic endoribonuclease activity. Successively, IRE leads to unconventional splicing of the XBP1 mRNA. The translation of this spliced transcription factor results in the transcriptional induction of genes expressing chaperones, ERAD components, and autophagy regulators. Previously, we have identified orthologues of IRE2, XBP1, and ATF6 in the genome of E. granulosus, the causative agent of human echinococcosis. In this work, we evolutionary and functionally characterized the IRE/XBP pathway in Echinococcus sp. Based on computational assays, we studied the evolutionary relationship of Echinococcus-IRE/XBP proteins among metazoans and performed a phylogenetic tree showing their position. Also, we showed that Echinococcus-IRE has considerable similarity in secondary and tertiary structures to human IRE (3p23.1A homodimer, 35% identity and 0.49 QMEANDisCo). Moreover, we determined that the level of total xbp1 transcript increased under ER-stress inducers treatment and, consequently provokes the enhancement of grp78 mRNA expression indicating the occurrence of ER stress in the parasite. Splicing of the xbp1 mRNA was analyzed to assess IRE1 activation. Through RT-PCR and sequencing, we corroborated the presence of unspliced- and spliced- xbp1 in the parasites. Additionally, since XBP-1 mediates the activation of TFEB, a transcription factor that promotes autophagy, we verified that IRE1/XBP1 activation induces autophagy, with overexpression of Echinococcus atg6, atg8 and tfeb genes. The crosstalk between the IRE1/XBP1 branch and autophagy highlights the importance of both mechanisms in parasite survival. Therefore, targeting the UPR-induced autophagy response may lead to novel therapeutic approaches.