title: Identification and cloning of a transglutaminase from giant freshwater prawn, Macrobrachium rosenbergii, and its transcription during pathogen infection and moulting.
abstract: Complementary (c)DNA encoding transglutaminase (TG) messenger (m)RNA of the giant freshwater prawn, Macrobrachium rosenbergii, was cloned from haemocytes by a reverse-transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) using oligonucleotide primers based on the TG sequence of the horseshoe crab, Tachypleus tridentatus; tiger shrimp, Penaeus monodon; kuruma shrimp, Marsupenaeus japonicus; and crayfish, Pacifastacus leniusculus. The 2722-bp cDNA contained an open reading frame (ORF) of 2334 bp, a 72-bp 5'-untranslated region (UTR), and a 316-bp 3'-UTR containing a stop codon and a poly A tail. The molecular mass of the deduced amino acid (aa) sequence (778 aa) was 86.67 kDa with an estimated pI of 5.4. The M. rosenbergii TG (abbreviated MrTG, accession no.: JF309296) contains a typical transglutaminase-like homologue, two putative integrin-binding motifs (RGD), ten glycosylation sites, and four calcium-binding sites; a catalytic triad is present as in arthropod TGs. Sequence comparison and a phylogenetic analysis revealed that shrimp TG can be separated into three subgroups, penaeid TG1, freshwater crustacean TG2 and marine crustacean TG2, and MrTG was more closely related to TG2 than to TG1. MrTG mRNA and TG activities were detected in all tested tissues of M. rosenbergii, with MrTG mainly being synthesised by haemocytes. There was a negative correlation between clotting time of haemolymph, and MrTG expression and TG activity of haemocytes in prawn injected with Lactococcus garvieae. The pattern of MrTG mRNA expression and TG activity in haemocytes exhibited a contrary tendency with clotting time of haemolymph during the moult stages. Those results indicate that cloned MrTG is involved in the defence response, and is probably the major functional TG for haemolymph coagulation in M. rosenbergii.
authors.affiliation: Department of Aquaculture, National Pingtung University of Science and Technology, Pingtung 91201, Taiwan, ROC.
authors.forename: Chun-Hung
authors.initials: CH
authors.lastname: Liu
authors.forename: Chin-Chyuan
authors.initials: CC
authors.lastname: Chang
authors.forename: Yun-Chih
authors.initials: YC
authors.lastname: Chiu
authors.forename: Winton
authors.initials: W
authors.lastname: Cheng
authors.forename: Maw-Sheng
authors.initials: MS
authors.lastname: Yeh
mesh_terms: D000595:Amino Acid Sequence; D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D001777:Blood Coagulation / Q000235:genetics*; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D040461:Gene Components; D006434:Hemocytes / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D018978:Molting / Q000235:genetics*; D016366:Open Reading Frames / Q000235:genetics; D033541:Palaemonidae / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D011503:Transglutaminases / Q000145:classification / Q000235:genetics* / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D011503:Transglutaminases