Chinese mitten crab (Eriocheir sinensis) iron-sulphur cluster assembly protein 2 (EsIscA2) is differentially regulated after immune and oxidative stress challenges

Abstract Iron-sulphur clusters (ISCs), one of the oldest and most versatile cofactors of proteins, are involved in catalysis reactions, electron transport reactions, regulation processes as well as sensing of ambient conditions. Iron-sulphur cluster assembly protein (IscA) is a scaffold protein member of ISC formation system, which plays a significant role in the assembly and maturation process of ISC proteins. In the present study, the cDNA sequence of iron-sulphur cluster assembly protein 2 (designated as Es IscA2) was cloned from Eriocheir sinensis . The open reading frame (ORF) of Es IscA2 was of 507 bp, encoding a peptide of 168 amino acids with a typically conserved Fe-S domain. A tetrameric form was predicated by the SWISS-MODEL prediction algorithm, and three conserved cysteine residues (Cys-93, Cys-158, Cys-160) from each IscA monomer were predicted to form a ‘cysteine pocket’. The deduced amino acid sequence of Es IscA2 shared over 50% similarity with that of other IscAs. Es IscA2 was clustered with IscA2 proteins from invertebrates and vertebrates, indicating that the protein was highly conservative in the evolution. r Es IscA2 exhibited a high iron binding affinity in the concentration ranging from 2 to 200 μM. Es IscA2 transcripts were detected in all the tested tissues including gonad, hemocytes, gill, muscle, heart, hepatopancreas and eyestalk, and Es IscA2 protein was detected in the mitochondria of hemocytes. The highest mRNA expression level of Es IscA2 was detected in muscle and hepatopancreas, which was about 34.66-fold ( p p Aeromonas hydrophila and lipopolysaccharide (LPS) stimulations, the mRNA expression of Es IscA2 in hemocytes was down-regulated and reached the lowest level at 24 h (0.31-fold, p p Es IscA2 mRNA in hepatopancreas was repressed from 6 h to 48 h post stimulation ( p 2 O 2 for 15 min, the expression level of Es IscA2 mRNA was significantly repressed to the 0.34–0.44-fold of that in the control group. After A. hydrophila stimulation, the mRNA expression of Es Grx2 was up-regulated at 3 h (3.22-fold compared to control group, p p Es IscA2 had iron-binding capabilities as observed in IscA proteins from other organisms, supporting the role of Es IscA2 as a mitochondrial iron donor for ISC synthesis in Chinese mitten crab. Its differential mRNA expression after immune and oxidative stress challenges suggested the adaptations of ISC synthesis rates to these stress conditions.