title: Three isoforms of anti-lipopolysaccharide factor identified from eyestalk cDNA library of swimming crab Portunus trituberculatus.
abstract: Anti-lipopolysaccharide factors (ALFs), as the potent antimicrobial peptides, can bind and neutralize lipopolysaccharide (LPS) and exhibit broad spectrum antimicrobial activities. In this study, three isoforms of the ALF homologues (PtesALF1-3) were identified from eyestalk cDNA library of swimming crab Portunus trituberculatus. The full-length cDNA sequences of PtesALF1, 2 and 3 were 1138, 1052 and 1057 bp encoding 92, 108 and 123 amino acids, respectively. PtesALF1-3 contained two conserved cysteine residues and shared high similarity with other reported ALFs. Predicted tertiary structures of PtesALF2 and 3 containing four β-strands and three α-helix were similar to that described in Limulus polyphemus, while PtesALF1 had only one α-helix in its spatial structure. Sequence analysis revealed PtesALF1-3 were encoded by the same genomic locus and generated by alternative splicing of the pre-mRNA. Totally 89 SNPs including 18 in coding region and 71 in noncoding region were detected by direct sequencing of 30 genomic samples. The mRNA expression of PtesALF1 and PtesALF1-3 transcripts was mainly detected in haemocytes but showed different expression pattern in other tissues including hepatopancreas, gill, eyestalk and muscle. After challenge with Vibrio alginolyticus, the temporal expression level of PtesALF1-3 transcripts in haemocytes showed a clear time-dependent response expression pattern with two peaks within the experimental period of 32 h, while PtesALF1 was up-regulated only once with obvious decrease at 6 h and significant increase at 24 h. These results suggest that the PtesALF isoforms have different tissue specificity and might provide multiple protective functions against invading bacteria in P. trituberculatus.
authors.affiliation: EMBL, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
authors.forename: Zhaoxia
authors.initials: Z
authors.lastname: Cui
authors.forename: Weisha
authors.initials: W
authors.lastname: Luan
authors.forename: Chengwen
authors.initials: C
authors.lastname: Song
authors.forename: Qing
authors.initials: Q
authors.lastname: Nie
authors.forename: Shuangyan
authors.initials: S
authors.lastname: Wang
authors.forename: Qianqian
authors.initials: Q
authors.lastname: Li
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003386:Brachyura / Q000145:classification / Q000235:genetics* / Q000276:immunology*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D015723:Gene Library; D007447:Invertebrate Hormones / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D010802:Phylogeny; D020033:Protein Isoforms; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D007447:Invertebrate Hormones; D020033:Protein Isoforms