title: A tailless Dscam from Eriocheir sinensis diversified by alternative splicing.
abstract: Dscam (Down syndrome cell adhesion molecule), a member of the immunoglobulin superfamily (IgSF), plays an essential role in pathogen recognition and further involves in the innate defense of invertebrates. In the present study, the cDNA of a Dscam from Chinese mitten crab Eriocheir sinensis (designated EsDscam) was cloned and characterized. It contained a 5-terminal untranslated region (UTR) of 60 bp, a 3-UTR of 216 bp with a poly (A) tail, and an open reading frame (ORF) of 4848 bp encoding a polypeptide of 1615 amino acids with the putative molecular mass of 178.4 kDa and theoretical isoelectric point of 6.31. The EsDscam protein shared higher sequence identities and similar domain architecture with Dscams from other invertebrate, including typical 10 immunoglobulin (Ig) domains, 6 ﬁbronectin type 3 domains (FNIII) and one cell attachment sequence (RGD) in extracellular region, while it lacked the expected transmembrane domain and cytoplasmic tail compared with other members of Dscam family. After sequencing 80 separate clones of Ig2, 3 and Ig7 regions from pooled cDNA libraries constructed from normal and bacterial-infected crabs, 44 alternative sequences were detected in the N-terminal of Ig2, 39 ones in Ig3, and 31 ones in Ig7 domain, suggesting that EsDscam could potentially encode at least 53196 unique isoforms. Furthermore, two 3'UTR isoforms and two 5'UTR isoforms of EsDscam were also identified by RACE strategy. EsDscam mRNA was most abundantly expressed in the tissues of nerve, muscle, hepatopancreas and gill, and weakly expressed in heart, gonad and hemocytes. Western blotting and immunofluorescence analysis revealed that EsDscam protein was mainly distributed in serum, and few on the membrane of crab hemocytes. These results suggested that this tailless EsDscam was one member of crustacean Dscam family, and the generation of diverse isoforms through alternative splicing allowed it to recognize various pathogens and play an active role in immune defense of crabs.
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China.
authors.forename: Jingjing
authors.initials: J
authors.lastname: Wang
authors.forename: Lingling
authors.initials: L
authors.lastname: Wang
authors.forename: Yang
authors.initials: Y
authors.lastname: Gao
authors.forename: Qiufen
authors.initials: Q
authors.lastname: Jiang
authors.forename: Qilin
authors.initials: Q
authors.lastname: Yi
authors.forename: Huan
authors.initials: H
authors.lastname: Zhang
authors.forename: Zhi
authors.initials: Z
authors.lastname: Zhou
authors.forename: Limei
authors.initials: L
authors.lastname: Qiu
authors.forename: Linsheng
authors.initials: L
authors.lastname: Song
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060829:Arthropod Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D015153:Blotting, Western / Q000662:veterinary; D003386:Brachyura / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005455:Fluorescent Antibody Technique / Q000662:veterinary; D006090:Gram-Negative Bacteria / Q000502:physiology; D006434:Hemocytes / Q000378:metabolism; D006458:Hemolymph / Q000378:metabolism; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010802:Phylogeny; D010843:Pichia / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment / Q000662:veterinary
chemical_list: D060829:Arthropod Proteins; D015815:Cell Adhesion Molecules; D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins
keywords: Alternative splicing; Dscam; Eriocheir sinensis; Immune defense