Avaliação da ativação de componentes da via de resposta a proteínas não dobradas (UPR) na infecção pelo Zika virus

Abstract

ABSTRACT Zika virus (ZIKV), of the genus Flavivirus, is the causative agent of an emerging infectious disease in many countries, with a high number of cases reported in recent years and a strong causal association with severe neurological syndromes. Understanding the behavior of the virus in the body is of paramount importance for the development of mechanisms to contain the disease, to conduct appropriate treatment and to introduce preventive measures. Viruses have evolved numerous strategies to manipulate host responses and ensure their replicative success. During viral replication there occurs high production and post-translational modifications of viral proteins in the endoplasmic reticulum (ER) of the host cells, which results in the overload of the organelle protein processing system and, consequently, activates an unfolded protein response pathway (UPR). This response is coordinated by the IRE1, PERK and ATF6 sensors coupled to the ER membrane which are responsible for monitoring gene transcription and translation of proteins involved in UPR control. This work addresses the effect of ZIKV infection on the activation of UPR pathway components in T98G human neuronal glioblastomas cells. The infection of these cells by ZIKV showed negative regulation of ATF6 in fluorescence microscopy and confocal microscopy assays, as well as the nonactivation of the transcriptional activity of ATF6 in a luciferase reporter gene assay. Furthermore, the expression of ATF6, XBP1, and CHOP-activated genes did not show elevation during ZIKV infection. The IRE1 sensor evaluated in the assay that monitored the processing of its substrate, the transcription factor mRNA XBP1, showed time-dependent modulation of infection, with down-regulation up to 48 hours post-infection. Expression of XBP1-activated genes, such as EDEM1, was activated within 72 hours of infection, suggesting late activation of this sensor. ZIKV infection induced lower gene expression of the PERK pathway, such as ATF4, CHOP, and GADD34, in the early stages of infection, ie activation of the expression of these genes occurred after 72 hours for Asian ZIKV. Already, an African sample of ZIKV assay induced earlier CHOP expression at 48 post-infection. Expression of 84 UPR pathway genes in infection with Asian ZIKV in a quantitative real-time PCR matrix assay generally increased significantly in later stages of infection. Evaluation of the production of BIP and calregulin chaperones was high only in infection with African ZIKV. Our findings showed that the ZIKV is capable of modulating UPR pathway components negatively, regulating ATF6 to late stages and IRE1 and PERK only in the early stages of infection, with the activation of the ER response pathway with greater intensity in the later phase of infection.