Comparative transcriptome analysis of the gills of Procambarus clarkii provides novel insights into the immune-related mechanism of copper stress tolerance

The red-swamp crayfish (Procambarus clarkii) is the most important economic shrimp species in China, and is an important model crustacean organism in many fields of research. In crustaceans, gills interface directly with the ambient environment and thus play a vital role in the toxicology. In the context of increasing environmental heavy metal pollution, the relationship between copper (Cu2+) stress and the immune response of P. clarkii has recently received considerable attention. However, impact of Cu2+ on the crayfish immune system is still not fully understood. In this study, we used Illumina sequencing technology to perform a transcriptome analysis of the gills of P. clarkii after 24 h of Cu2+ treatment. A total of 37,226,812 unigenes were assembled, and 1943 unigenes were significantly differentially expressed between the control and Cu2+ treatment groups. Functional categorization of differentially expressed genes (DEGs) revealed that genes related to antioxidant activity, detoxication, metabolic processes, biosynthetic processes, and immune system processes were differentially regulated during Cu2+ stress. In addition, DEGs in the immune system were classified as being related to the MAPK signaling pathway, purine metabolism, Toll and Imd signaling pathway, PI3K-Akt signaling pathway and Hippo signaling pathway. Five genes (CuZnSOD, CAT, IDH1, PHYH and DECR2) were significantly up-regulated in the peroxisome pathway, which plays an important role in reacting to oxidative stress. Importantly, qRT-PCR validation of the results for seven genes chosen at random (NDK, ATP6L, ATP5C1, RPS14, RPL22e, CTSF and HSP90A) confirmed the Illumina sequencing results. This study provides a valuable starting point for further studies to elucidate the molecular basis of the immune system's response to Cu2+ stress in crayfish.