The mannose-binding lectin (MBL) gene cloned from Exopalaemon carinicauda plays a key role in resisting infection by Vibrio parahaemolyticus.

Mannose-binding lectin (MBL) is a vital member of the lectin family, crucial for mediating functions within the complement lectin pathway. In this study, following the cloning of the mannose-binding lectin (MBL) gene in the ridgetail white prawn, Exopalaemon carinicauda, we examined its expression patterns across various tissues and its role in combating challenges posed by Vibrio parahaemolyticus. The results revealed that the MBL gene spans 1342 bp, featuring an open reading frame of 972 bp. It encodes a protein comprising 323 amino acids, with a predicted relative molecular weight of 36 kDa and a theoretical isoelectric point of 6.18. The gene exhibited expression across various tissues including the eyestalk, heart, gill, hepatopancreas, stomach, intestine, ventral nerve cord, muscle, and hemolymph, with the highest expression detected in the hepatopancreas. Upon challenge with V. parahaemolyticus, RT-PCR analysis revealed a trend of MBL expression in hepatopancreatic tissues, characterized by an initial increase followed by a subsequent decrease, peaking at 24 h post-infection. Employing RNA interference to disrupt MBL gene expression resulted in a significant increase in mortality rates among individuals challenged with V. parahaemolyticus. Furthermore, we successfully generated the Pet32a-MBL recombinant protein through the construction of a prokaryotic expression vector for conducting in vitro bacterial inhibition assays, which demonstrated the inhibitory effect of the recombinant protein on V. parahaemolyticus, laying a foundation for further exploration into its immune mechanism in response to V. parahaemolyticus challenges.

Expression and function analysis of crustacyanin gene family involved in resistance to heavy metal stress and body color formation in Exopalaemon carinicauda.

Crustacyanin has the function of binding astaxanthin which is the best antioxidant, and plays an important role in the body color variation of crustaceans. To investigate the causes of body color variation of the ridgetail white prawn, Exopalaemon carinicauda, the present study obtained four subtypes of crustacyanin gene: C1, C2, A1, and A2. Based on fluorescence quantitative polymerase chain reaction, lipocalin-C1 is mainly expressed in the eyestalk, lipocalin-C2 is in the ventral nerve cord, and lipocalin-A1 and lipocalin-A2 are in subcutaneous adipose tissues. Under the inhibiting effect of Cd2+ stress, the expression of four subtypes first increases and then decreases within 24 h, and reaches the maximum at 6 or 12 h. RNA interference experiments showed a decrease in the expression of lipocalin genes in subcutaneous adipose tissue for each subtype, with the body color changing from transparent to red, and the dark red spots on the epidermis changing to bright red. Moreover, the blue protein in the subcutaneous adipose tissue largely disappeared, based on the light micrographs. In view of these findings, the crustacyanin gene appears to fulfill some function in the resistance to heavy metal stress and body color formation of E. carinicauda.

Cloning and functional study of lipocalin: retinol-binding protein-like gene family of the ridgetail white prawn, Exopalaemon carinicauda.

Lipocalin is a large family with complex functions including retinol-binding protein (RBP), crustacyanin (CRCN), apolipoprotein D, etc. In shrimps, it is well known that CRCN is related to body color. Recently, retinoic acid/retinol-binding protein was found in shrimp. However, little is known about the function of RBP and relationships among the gene members of lipocalin in shrimps. Based on the transcriptome sequences responding to starvation stress, three genes of the lipocalin-retinol-binding protein-like gene family (lipocalin-1, lipocalin-2, and lipocalin-3) were cloned by RACE from the ridgetail white prawn, Exopalaemon carinicauda. Homology analysis showed that these three genes had high similarity with the known insect apolipoprotein D gene and vertebrate retinol-binding protein gene, and they are of the same type in terms of evolution. Fluorescence quantitative PCR showed that the above three genes were mainly expressed in the ventral nerve cord of E. carinicauda. The expression characteristics of the three genes at different developmental stages showed that they were more highly expressed at the larval stage, which suggests that they might be related to embryonic and larval development. The RNA interference tests showed that after silencing lipocalin-1 and lipocalin-3, the body color of individual shrimps turned slightly red and the blue pigment in the epidermis largely disappeared, but no significant change took place in the appearance of individuals after silencing lipocalin-2. In addition, on the 6th and 16th days of interference, dead shrimps appeared in the lipocalin-1 and lipocalin-3 interference groups. The dead shrimps had hard crusts and remained in a molting posture. Totally, this study showed that the retinol-binding protein-like gene obtained in this study had certain biological functions in the growth and development and body color formation as CRCN; in addition, it also plays a role in nerve system and molting of E. carinicauda.