Abstract:Exopalaemon modestus is one of the main shrimps which has high economic value.But its genetic information and functional gene background are rarely known so far.In this study,the cDNAs of four novel C-type lectins(designated as EmCTL-1,EmCTL-2, EmCTL-3 and EmCTL-4)were cloned from the ovary of white shrimp,E.modestus,by means of PCR and 5′and 3′rapid amplification of cDNA ends (RACE).The full-length cDNAs consist of 1 390,1 119,1 364 and 1 299 bp with one open reading frame consisting of 996,969,1 026 and 1 044 bp,encoding 332,322,341 and 347 amino acids respectively.Their deduced amino acid sequences contained a putative signal peptide of 19,21,20 and 17 amino acid residues respectively.The four C-type lectin genes all contained two carbohydrate recognition domains,CRD1 and CRD2.The CRD1 was composed of 128 residues in EmCTL-2 and EmCTL-3,127 residues in EmCTL-1 and 126 residues in EmCTL-4. And the CRD2 was composed of 127 residues in EmCTL-1 and EmCTL-3,129 residues in EmCTL-2 and 130 residues in EmCTL-4.Both the CRD1 and CRD2 had 6 conserved cysteine residues and 3 conserved tryptophane residues.The motif of Ca2 binding sites 2 were EPA and FND in EmCTL-1,EPA and FHD in EmCTL-2,WHD and FDD in EmCTL-3 and EPD and FRD in EmCTL-4.So EmCTL-1,EmCTL-2 and EmCTL-4 can be deduced binding with mannose in immune reactions.And the four C-type lectin genes shared 40%,37%,69% and 73% identity with the parallel C-type lectin genes in Macrobrachium rosenbergii,existing in Palaemonidae.The CODEML program implemented in the PAML package was used to test for variations in selective pressures of shrimp and prawns C-type lectin genes with two CRDs,included the four genes acquired in this study.The result showed that the shrime and prawns lectin genes with two CRDs had undergone positive selection.According to the LTR test,3.8% of the sites were driven by positive selection in the M2a model,while 5.6% of the sites were driven by positive selection in the M8 model.Phylogenetic analysis showed that the lectins from the decapods could be divided into four clear clusters.Cluster C included the dual-CRD lectins while cluster D includ the single-CRD lectins,and cluster A included the dual-CRD lectins except the PmCTL-1 in tiger shrimp,cluster B included the single-CRD lectins except the FcCTL in Chinese shrimp.It indicated that the numbers of CRD played a key role in phyletic evolution.The results showed that EmCTL-1,EmCTL-2, EmCTL-3 belonged to cluster C and EmCTL-4 belonged to cluster A.Four novel C-type lectin genes in E.modestus had a close evolutionary relationship to the C-type lectins of giant freshwater prawn.Tissue distribution analysis using semi-quantitative RT-PCR methods showed that EmCTL-1 was main expressed in hemolymph,testis and ovary.EmCTL-3 and EmCTL-4 could be detected main in testis,ovary,hepatopancreas,hemolymph,heart and thoracic ganglion while EmCTL-2 was mainly expressed in ovary,testis,hemolymph,thoracic ganglion and heart.The results showed that the hemolymph was the mainly expression tissue of four C-type lectin genes in E.modestus.Interestingly,and the testis was the organ with highest expression level of C-type lectin transcripts in this study.It can be speculated that the C-type lectins in E.modestus may agglutinate microorganisms in the hemolymph for possessing signal peptides,conserved cysteine residues,conserved tryptophane residues,high expression level in hemolymph and observed positive selection.