Characterization, Stress Response and Functional Analyses of Giant River Prawn (Macrobrachium rosenbergii) Glucose-Regulated Protein 78 (Mr-grp78) under Temperature Stress and during Aeromonas hydrophila Infection.

Abstract

Simple SummaryGlucose-regulated protein 78 (grp78) is classified as a member of the Hsp70 subfamily. This protein functions as a key factor in signal transduction associated with the unfolded protein response (UPR) in the endoplasmic reticulum (ER) during cellular stress and protects against cell damage caused by toxic chemicals, oxidative stress, Ca2+ depletion, programmed cell death and various infectious conditions. To investigate this crucial mechanism in giant river prawn (Macrobrachium rosenbergii), we analyzed the biological function of prawn grp78 at the molecular level in this study. The regulation of this gene was intensively analyzed under normal bacterial infection and heat/cold-shock inductions. A functional analysis of this gene under heat and infectious stress conditions was performed by gene knockdown. The information obtained in the current study clearly indicates the crucially significant roles of grp78 in the cellular stress responses of the target experimental animal under various stress conditions.The endoplasmic reticulum (ER) is an organelle important for several functions of cellular physiology. This study identified the giant river prawn’s glucose-regulated protein 78 (Mr-grp78), which is important for ER stress mechanisms. Nucleotide and amino acid analyses of Mr-grp78, as compared with other species, revealed the highest similarity scores with the grp78 genes of crustaceans. An expression analysis by quantitative RT-PCR indicated that Mr-grp78 was expressed in all tissues and presented its highest expression in the ovary (57.64 ± 2.39-fold), followed by the gills (42.25 ± 1.12), hindgut (37.15 ± 2.47), thoracic ganglia (28.55 ± 2.45) and hemocytes (28.45 ± 2.26). Expression analysis of Mr-grp78 mRNA levels under Aeromonas hydrophila induction and heat/cold-shock exposure was conducted in the gills, hepatopancreas and hemocytes. The expression levels of Mr-grp78 in these tissues were highly upregulated 12 h after bacterial infection. In contrast, under heat- and cold-shock conditions, the expression of Mr-grp78 was significantly suppressed in the gills at 24–96 h and in the hepatopancreas at 12 h (p < 0.05). A functional analysis via Mr-grp78 gene knockdown showed that Mr-grp78 transcription in the gills, hepatopancreas and muscle strongly decreased from 6 to 96 h. Furthermore, the silencing of this gene effectively increased the sensitivity of the tested prawns to heat- and pathogenic-bacterium-induced stress. The results of this study clearly demonstrate the significant functional roles of Mr-grp78 in response to both temperature and pathogen treatments.