title: Cathepsin L of the sea cucumber Apostichopus japonicus-molecular characterization and transcriptional response to Vibrio splendidus infection.
abstract: Cathepsin L, a lysosomal endopeptidase, has been noted for its involvement in the innate immune response in invertebrates. Here, the cathepsin L cDNA of the sea cucumber Apostichopus japonicus (AjCatL) is identified from an EST library and then cloned by the rapid amplification of the cDNA ends (RACE) PCR. The full-length cDNA is 1678 bp long containing an open reading frame (ORF) of 1002 bp, an 80 bp 5' UTR and a 599 bp 3' UTR. The cDNA encodes 333 amino acid residues with a predicted molecular mass of 37.07 kDa and a theoretical isoelectric point (pI) of 5.01. The full-length AjCatL contains three active sites of eukaryotic thiol (cysteine) protease at positions 133-144, 278-288 and 295-314. Analysis of the predicted tertiary structure of prepro-CatL (17-333 aa) and mature-CatL (116-333 aa) reveals that the propeptide region (17-115 aa) blocks access to the substrate-binding cleft. Phylogenetic analysis shows that the AjCatL is clustered together with two other CatLs from Strongylocentrotus purpuratus. The enzymatic activity of AjCatL was verified using a substrate hydrolyzing assay with recombinant mAjCatL. Further analysis of real time-PCR demonstrates that the expression of AjCatL mRNA is significantly up-regulated in the coelomocytes in cases of infection with the common bacterial pathogen, Vibrio splendidus. This suggests that the AjCatL is likely to be involved in the immune response.
authors.affiliation: National Engineering Research Center of Marine Facilities Aquaculture, College of Marine Sciences, Zhejiang Ocean University, Zhoushan, Zhejiang, 316022, China.
authors.forename: Jingwen
authors.initials: J
authors.lastname: Yang
authors.affiliation: National Engineering Research Center of Marine Facilities Aquaculture, College of Marine Sciences, Zhejiang Ocean University, Zhoushan, Zhejiang, 316022, China.
authors.forename: Huihui
authors.initials: H
authors.lastname: Liu
authors.affiliation: Ocean Research Center of Zhoushan, Zhejiang University, Zhoushan, Zhejiang, 316021, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Zheng
authors.affiliation: National Engineering Research Center of Marine Facilities Aquaculture, College of Marine Sciences, Zhejiang Ocean University, Zhoushan, Zhejiang, 316022, China.
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Xiang
authors.affiliation: National Engineering Research Center of Marine Facilities Aquaculture, College of Marine Sciences, Zhejiang Ocean University, Zhoushan, Zhejiang, 316022, China. Electronic address: nblzmnb@163.com.
authors.forename: Zhenming
authors.initials: Z
authors.lastname: Lv
authors.affiliation: National Engineering Research Center of Marine Facilities Aquaculture, College of Marine Sciences, Zhejiang Ocean University, Zhoushan, Zhejiang, 316022, China. Electronic address: wangtianming@zjou.edu.cn.
authors.forename: Tianming
authors.initials: T
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D056668:Cathepsin L / Q000235:genetics* / Q000276:immunology*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D007113:Immunity, Innate; D008969:Molecular Sequence Data; D010802:Phylogeny; D012333:RNA, Messenger / Q000235:genetics; D016415:Sequence Alignment; D047528:Stichopus / Q000145:classification / Q000235:genetics* / Q000276:immunology / Q000382:microbiology*; D015854:Up-Regulation; D014733:Vibrio / Q000502:physiology*
chemical_list: D018076:DNA, Complementary; D012333:RNA, Messenger; D056668:Cathepsin L
keywords: Apostichopus japonicus; Cathepsin L; Gene expression; Immune response; Vibrio splendidus