Molecular cloning and characterization of lipopolysaccharide- and beta-1,3-glucan-binding protein from the giant freshwater prawn Macrobrachium rosenbergii and its transcription in relation to foreign material injection and the molt stage.
Lipopolysaccharide (LPS)- and beta-1,3-glucan-binding protein (LGBP) complementary (c)DNA was cloned from the hepatopancreas of the giant freshwater prawn Macrobrachium rosenbergii using oligonucleotide primers and a reverse-transcription polymerase chain reaction (RT-PCR). Both 3'- and 5'-regions were isolated by the rapid amplification of cDNA ends (RACE) method. Analysis of the nucleotide sequence revealed that the cDNA clone has an open reading frame of 1389 bp encoding a protein of 378 amino acids (aa) including a 15-aa signal peptide. The calculated molecular mass of the mature protein (363 aa) was 41.2 kDa with an estimated pI of 4.73. The M. rosenbergii LGBP sequence contains (1) three putative integrin-binding motifs, (2) a glucanase motif, (3) a putative N-glycosylation site, (4) four protein kinase C phosphorylation sites, (5) four casein kinase II phosphorylation sites, and (6) a putative recognition motif. Sequence comparison showed that the deduced amino acids of LGBP of M. rosenbergii had overall similarities of 60-71% to those of known crustacean LGBPs and beta-1,3-glucan-binding proteins (BGBPs). The LGBP of M. rosenbergii was mainly expressed in the hepatopancreas. The LGBP transcript of M. rosenbergii was downregulated in haemocytes, but was upregulated in the hepatopancreas when injected with LPS and poly:IC after 12 h. The LGBP messenger (m)RNA expression of prawns in the postmolt stage was significantly upregulated in haemocytes, but downregulated in the hepatopancreas, which revealed a complementary relationship between haemocytes and the hepatopancreas in the molt cycle.
Yeh
Maw-Sheng
MS
Institute of Biomedical Nutrition, Hungkuang University, Sha-Lu, Taichung 43302, Taiwan, ROC.
Chang
Chin-Chyuan
CC
Cheng
Winton
W
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D043143:Hepatopancreas / Q000276:immunology*; D037102:Lectins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D018978:Molting / Q000276:immunology; D033541:Palaemonidae / Q000235:genetics / Q000276:immunology*; D010802:Phylogeny; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique; D016415:Sequence Alignment; D014158:Transcription, Genetic
D002352:Carrier Proteins; D037102:Lectins; D012333:RNA, Messenger; C037937:glucan-binding proteins