Cloning, Functional Characterization and Response to Cadmium Stress of the Thioredoxin-like Protein 1 Gene from Phascolosoma esculenta.

Jiajie Meng,Congcong Hou,Xinming Gao,Jianping Wang,Chundan Zhang,Shengyu Luo,Chen Du,Chenwen Lin,Junquan Zhu,Daojun Tang,Shan Jin

doi:10.3390/ijms23010332

Abstract

Cadmium (Cd) is a heavy metal toxicant and is widely distributed in aquatic environments. It can cause excessive production of reactive oxygen species (ROS) in the organism, which in turn leads to a series of oxidative damages. Thioredoxin (Trx), a highly conserved disulfide reductase, plays an important role in maintaining the intracellular redox homeostasis in eukaryotes and prokaryotes. Phascolosoma esculenta is an edible marine worm, an invertebrate that is extensively found on the mudflats of coastal China. To explore the molecular response of Trx in mudflat organisms under Cd stress, we identified a new Trx isoform (Trx-like protein 1 gene) from P. esculenta for the first time, designated as PeTrxl. Molecular and structural characterization, as well as multiple sequence and phylogenetic tree analysis, demonstrated that PeTrxl belongs to the Trx superfamily. PeTrxl transcripts were found to be ubiquitous in all tissues, and the highest expression level occurred in the coelomic fluid. Exposure to three sublethal concentrations of Cd resulted in the upregulation and then downregulation of PeTrxl expression levels over time in coelomic fluid of P. esculenta. The significant elevation of PeTrxl expression after 12 and 24 h of Cd exposure at 6 and 96 mg/L, respectively, might reflect its important role in the resistance to Cd stress. Recombinant PeTrxl (rPeTrxl) showed prominent dose-dependent insulin-reducing and ABTS free radical-scavenging abilities. After exposure to 96 mg/L Cd for 24 h, the ROS level increased significantly in the coelomic fluid, suggesting that Cd induced oxidative stress in P. esculenta. Furthermore, the injection of rPeTrxl during Cd exposure significantly reduced the ROS in the coelomic fluid. Our data suggest that PeTrxl has significant antioxidant capacity and can protect P. esculenta from Cd-induced oxidative stress.

Highlights

Cadmium (Cd), an unessential metal element in organisms, is widely distributed in aquatic environments [1]
This study indicated that PeTrxl is involved in the scavenging of reactive oxygen species (ROS) induced by Cd and provides important insights into the role of antioxidant systems during heavy metal-induced oxidative stress resistance
The results showed that in the presence of dithiothreitol (DTT), the disulfide bonds between the A and B chains of insulin were broken in the experimental group with Recombinant PeTrxl (rPeTrxl), with the precipitation caused by B chain aggregation increasing the absorbance (Ab) of the reaction system at 650 nm

Summary

Introduction

Cadmium (Cd), an unessential metal element in organisms, is widely distributed in aquatic environments [1]. Cd2+ in organisms can bind with the sulfhydryl groups of antioxidant enzymes, resulting in their reduced activity or inactivation and indirectly increasing intracellular levels of reactive oxygen species (ROS) [12,13]. Generating OH radicals via the Fenton reaction between Cd2+ and hydrogen peroxide directly leads to an increase in intracellular levels of ROS [14]. The excessive ROS can cause lipid peroxidation of the cell membrane, leading to oxidative damage or apoptosis [15,16]. To minimize the adverse effects of ROS, developing pathways through which excess ROS are scavenged is necessary for organisms

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Discussion

Conclusion