Molecular Characterization and Expression Analysis of Glutaredoxin 5 in Black Tiger Shrimp (Penaeus monodon) and Correlation Analysis Between the SNPs of PmGrx5 and Ammonia-N Stress Tolerance Trait

Abstract

Glutaredoxins (Grxs) are glutathione-dependent oxidoreductases that belong to the thioredoxin (Trx) superfamily and are an essential part of the redox system in living organisms. However, there is a serious lack of sequence information and functional validation associated with Grxs in crustaceans. In this study, a new Grx gene (PmGrx5) was identified and characterized in black tiger shrimp (Penaeus monodon). The full-length cDNA of PmGrx5 is 787 bp and consists of 114 bp 5′-UTR, 232 bp 3′-UTR, and 441 bp ORF, encoding a hypothetical protein of 146 amino acids. The putative PmGrx5 protein is 16.27 kDa with a theoretical isoelectric point of 5.90. Sequence alignment showed that PmGrx5 had the highest amino acid sequence homology with Grx5 from Penaeus vannamei at 98.63% and clustered with Grx5 from other crustaceans. Quantitative real-time PCR (qRT-PCR) analysis showed that PmGrx5 was expressed in all tissues examined, with a higher expression in the testis, stomach, lymphoid organ, and gill. PmGrx5 was continuously expressed during development, with the highest expression in zoea I. Ammonia-N stress and bacterial infection both differentially upregulated PmGrx5 expression in the hepatopancreas and gill. In addition, when PmGrx5 was inhibited, the expression of some other antioxidant enzymes was upregulated at the beginning of ammonia-N stress, but as the stress time increased, the expression of antioxidant enzymes was inhibited, the expression of apoptotic genes was increased, and the GSH content was significantly reduced. Inhibition of PmGrx5 led to a greater risk of oxidative damage in shrimp. In addition, the relationship between SNPs in exons of the PmGrx5 gene and tolerance to ammonia-N stress was identified and analysed. A total of nine SNPs were successfully identified, eight of which were significantly associated with ammonia and nitrogen stress tolerance trait in shrimp (P < 0.05). The present study shows that PmGrx5 is involved in redox regulation and plays an important role in shrimp resistance to marine environmental stresses. Meanwhile, this study will provide a basis for molecular marker breeding in shrimp.