Identification and functional characterization of an endoplasmic reticulum oxidoreductin 1-α gene in Litopenaeus vannamei

Abstract

In the current study, full-length sequence of endoplasmic reticulum oxidoreductin 1-α (LvERO1-α) was cloned from Litopenaeus vannamei. Real-time RT-PCR results showed that LvERO1-α was highly expressed in hemocytes, gills, and intestines. White spot syndrome virus (WSSV) challenge was performed, and the expression of LvERO1-α and two other downstream genes of the double-stranded RNA-activated protein kinase-like ER kinase-eIF2α (PERK-α) pathway, namely, homocysteine-induced endoplasmic reticulum protein (LvHERP) and acylamino-acid-releasing enzyme (LvAARE), strongly increased in the hemocytes. Flow cytometry assay results indicated that the apoptosis rate of L. vannamei hemocytes in the LvERO1-α knockdown group was significantly lower than that of the controls. Moreover, shrimps with knockdown expression of LvERO1-α exhibited decreased cumulative mortality upon WSSV infection. Downregulation of L. vannamei immunoglobulin-binding protein (LvBip), which had been proven to induce unfolded protein response (UPR) in L. vannamei, did not only upregulate LvERO1-α, LvHERP, and LvAARE in hemocytes, but also increased their apoptosis rate, as well as the shrimp cumulative mortality. Furthermore, reporter gene assay results showed that the promoter of LvERO1-α was activated by L. vannamei activating transcription factor 4, thereby confirming that LvERO1-α was regulated by the PERK-eIF2α pathway. These results suggested that LvERO1-α plays a critical role in WSSV-induced apoptosis, which likely occurs through the WSSV-activated PERK-eIF2α pathway.