Role of transglutaminase in immune defense against bacterial pathogens via regulation of antimicrobial peptides

• A 3248-bp (full-length) TGase cDNA in Eriocheir sinensis was cloned, with a 2274-bp ORF encoding a 757 amino acid protein.
• EsTGase was mainly expression in hemocytes and up-regulated post-challenge with V. parahaemolyticus and S. aureus.
• The activity of EsTGase was significantly decrease post V. parahaemolyticus and S. aureus challenging respectively.
• Knockdown of EsTGase down-regulated expression of immune-related genes with or without bacteria stimulation in vitro.
• Absence of EsTGase led to higher bacterial counts in the hemocyte culture medium.

Transglutaminase (TGase) is critical for blood coagulation, a conserved immunological defense mechanism among invertebrates. Here, a 3248-bp (full-length) TGase cDNA in Eriocheir sinensis (EsTGase) was cloned, with a 2274-bp open reading frame (ORF) encoding a 757 amino acid protein containing two transglut domains, one TGase/protease-like homolog domain and a KGD (Lys-Gly-Asp) motif. Phylogenetic analysis demonstrated that EsTGase appeared earlier in evolution compared with TGases of other crustaceans and mammals. EsTGase mRNA was mainly detected in hemocytes and up-regulated post-challenge with bacteria (Vibrio parahaemolyticus and Staphylococcus aureus), suggesting an immune function for this gene. Moreover, the EsTGase activity in hemocytes challenged with V. parahaemolyticus and S. aureus was decreased significantly. RNA interference of EsTGase down-regulated expression of immune-related genes CrusEs2, EsLecG and Es-DWD1 with or without bacteria stimulation in vitro. Furthermore, absence of EsTGase led to higher bacterial counts in the hemocyte culture medium. Thus, EsTGase is an important component of the crab immune response and is involved in the regulation of certain immune-related genes, particularly those encoding anti-microbial peptides.