Molecular cloning and expression of NOS in shrimp, Litopenaeus vannamei.

The importance of the nitric oxide synthase (NOS) gene family is demonstrated by many studies in recent years. However, the lack of sequence information and clones of shrimp NOS cDNA limits further study on its characterization and function in this species. In this report, the cDNA of NOS contained full-length ORF was cloned from the Pacific white shrimp, Litopenaeus vannamei. It was of 4680 bp, including a 5'-terminal untranslated region (UTR) of 278 bp, a 3'-terminal UTR of 862 bp, which contained 5 ATTTA repeats, and an open reading frame (ORF) of 3540 bp encoding a polypeptide of 1179 amino acids. It contained a typical NO synthase domain at the N-terminal, next to a flavodoxin 1 domain, a flavin adenine dinucleotide (FAD) binding domain, respectively, and a conservative nicotinamide adenine dinucleotide (NAD) binding domain structure at the C-terminal. Quantitative real-time reverse transcription PCR analysis revealed L. vannamei NOS (LvNOS) to be expressed in most shrimp tissues, with highest expression in the hepatopancreas and weakest expression in skin. The expression of LvNOS after challenge with LPS and poly I:C was tested in hemocytes, hepatopancreas and nerve. The results indicated that the NOS transcript level could be induced in hemocytes by injection with LPS. The highest expression was in the hemocyte, with 8.8 times (at 3 h) as much as that in the control (p < 0.05). However, sharp down-regulation of NOS was found in hepatopancreas and nerve after LPS and poly I:C injection (p < 0.05). These results suggested that NOS might play an important role in shrimp's defense against pathogenic infection.

Yao CL ，Ji PF ，Wang ZY ，Li FH ，Xiang JH ... -  《-》

Molecular cloning and expression profiles of nitric oxide synthase (NOS) in mud crab Scylla paramamosain.

Li S ，Zhang Z ，Li C ，Zhou L ，Liu W ，Li Y ，Zhang Y ，Zheng H ，Wen X ... -  《-》

Two types of calmodulin play different roles in Pacific white shrimp (Litopenaeus vannamei) defenses against Vibrio parahaemolyticus and WSSV infection.

Calmodulin (CaM) plays an important role in calcium-dependent signal transduction pathways. In the present study, two alternative splicing isoforms of CaM (named LvCaM-A and LvCaM-B) cDNA were cloned from the Pacific white shrimp, Litopenaeus vannamei. LvCaM-A was of 1101 bp, including a 5'-terminal untranslated region (UTR) of 70 bp, a 3'-terminal UTR of 581 bp and an open reading frame (ORF) of 450 bp encoding a polypeptide of 149 amino acids with a calculated molecular weight (Mw) of 17 kDa and pI of 4.41. LvCaM-B was 689 bp, including a same 5'-UTR of 70 bp, a 3'-terminal UTR of 109 bp and an ORF of 510 bp encoding a polypeptide of 169 amino acids with a calculated Mw of 19 kDa and pI of 4.36. Both LvCaM-A and LvCaM-B contained 4 conservative EF-hand motifs. Quantitative real-time reverse transcription PCR analysis revealed LvCaM-A to be expressed in most shrimp tissues, with the predominant expression in nerve and the weakest expression in heart. However, LvCaM-B expression level was much weaker than those of LvCaM-A in all the tested tissues with main expression in hepatopancreas. The expression of LvCaM-A and LvCaM-B after challenge with Vibrio parahaemolyticus and WSSV were tested in hemocytes, hepatopancreas and nerve. The results indicated that LvCaM-A and LvCaM-B transcripts could be significantly induced in hemocytes and hepatopancreas respectively by injection with V. parahaemolyticus. The highest expression of LvCaM-A was in the hemocytes with 2.3 times (at 48 h) greater expression than in the control (p < 0.05). However, sharp down-regulation of both LvCaM-A and LvCaM-B were detected in nerve after Vibrio- and WSSV injection (p < 0.05). These results suggested that CaM might play an important role in shrimp's defense against pathogenic infection.

Ji PF ，Yao CL ，Wang ZY 《-》

cDNA cloning, identification, tissue localisation, and transcription profile of a transglutaminase from white shrimp, Litopenaeus vannamei, after infection by Vibrio alginolyticus.

Yeh MS ，Liu CH ，Hung CW ，Cheng W ... -  《-》

Identification of fatty acid synthase from the Pacific white shrimp, Litopenaeus vannamei and its specific expression profiles during white spot syndrome virus infection.

Fatty acid synthase (FAS) in animal tissues consists of two identical monomers and is known to be a complex multi-functional enzyme that plays an important role in energy homeostasis. However, there are few reports of studies focused on the relationship between FAS and virus infection in invertebrates. In the present study, we cloned the FAS gene from an economically important invertebrate, the Pacific white shrimp Litopenaeus vannamei. The full-length FAS cDNA is 8268 bp, including a 5'-terminal untranslated region of 137 bp, a 3'-terminal untranslated region of 601 bp and an open reading frame of 7530 bp. FAS cDNA encodes a polypeptide of 2509 amino acid residues that contains a typical β-ketoacyl synthase (KS) domain at the N-terminus, next to a malonyl/acetyltransferase (MAT) domain, a dehydrase domain, an enoyl reductase domain, a ketoacyl reductase domain, a phosphopantetheine attachment site domain and a thioesterase domain at the C-terminus. Quantitative real-time RT-PCR revealed the up-regulated expression of FAS in L. vannamei hepatopancreas and muscle after white spot syndrome virus (WSSV) infection. The expression of FAS in muscle was 13.03-fold greater than that in the control (p<0.05) and 2.93-fold greater in hepatopancreas (p>0.05). Meanwhile, expression of the fatty acid-binding protein (FABP), another important factor in lipid metabolism, was increased in muscle to 19.20-fold greater than that in the control (p<0.05) but only 0.76-fold in hepatopancreas (p>0.05). These results implied that WSSV infected body surface tissues, but there was very little infection of internal organs. We suggest that the increase of FAS expression is induced in WSSV-infected shrimps, and the virus changes the lipid metabolism of the host, which directly affects virus assembly or defense against virus infection.

Yang L ，Liu J ，Liu M ，Qian M ，Zhang M ，Hu H ... -  《-》