title: Identification, characterization, and functional analysis of Tube and Pelle homologs in the mud crab Scylla paramamosain.
abstract: Tube and Pelle are essential components in Drosophila Toll signaling pathway. In this study, we characterized a pair of crustacean homologs of Tube and Pelle in Scylla paramamosain, namely, SpTube and SpPelle, and analyzed their immune functions. The full-length cDNA of SpTube had 2052 bp with a 1578 bp open reading frame (ORF) encoding a protein with 525 aa. A death domain (DD) and a kinase domain were predicted in the deduced protein. The full-length cDNA of SpPelle had 3825 bp with a 3420 bp ORF encoding a protein with 1140 aa. The protein contained a DD and a kinase domain. Two conserved repeat motifs previously called Tube repeat motifs present only in insect Tube or Tube-like sequences were found between these two domains. Alignments and structure predictions demonstrated that SpTubeDD and SpPelleDD significantly differed in sequence and 3D structure. Similar to TubeDD, SpTubeDD contained three common conserved residues (R, K, and R) on one surface that may mediate SpMyD88 binding and two common residues (A and A) on the other surface that may contribute to Pelle binding. By contrast, SpPelleDD lacked similar conservative residues. SpTube, insect Tube-like kinases, and human IRAK4 were found to be RD kinases with an RD dipeptide in the kinase domain. SpPelle, Pelle, insect Pelle-like kinases, and human IRAK1 were found to be non-RD kinases lacking an RD dipeptide. Both SpTube and SpPelle were highly expressed in hemocytes, gills, and hepatopancreas. Upon challenge, SpTube and SpPele were significantly increased in hemocytes by Gram-negative or Gram-positive bacteria, whereas only SpPelle was elevated by White Spot Syndrome Virus. The pull-down assay showed that SpTube can bind to both SpMyD88 and SpPelle. These results suggest that SpTube, SpPelle, and SpMyD88 may form a trimeric complex involved in the immunity of mud crabs against both Gram-negative and Gram-positive bacteria.
authors.affiliation: Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation and Utilization, Ministry of Agriculture, Shanghai, P. R. China ; East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai, P. R. China.
authors.forename: Xin-Cang
authors.initials: XC
authors.lastname: Li
authors.forename: Xiao-Wen
authors.initials: XW
authors.lastname: Zhang
authors.forename: Jun-Fang
authors.initials: JF
authors.lastname: Zhou
authors.forename: Hong-Yu
authors.initials: HY
authors.lastname: Ma
authors.forename: Zhi-Dong
authors.initials: ZD
authors.lastname: Liu
authors.forename: Lei
authors.initials: L
authors.lastname: Zhu
authors.forename: Xiao-Juan
authors.initials: XJ
authors.lastname: Yao
authors.forename: Lin-Gui
authors.initials: LG
authors.lastname: Li
authors.forename: Wen-Hong
authors.initials: WH
authors.lastname: Fang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060829:Arthropod Proteins / Q000737:chemistry / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003386:Brachyura / Q000235:genetics* / Q000382:microbiology / Q000821:virology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D006458:Hemolymph / Q000378:metabolism* / Q000382:microbiology / Q000821:virology; D054884:Host-Pathogen Interactions; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000502:physiology; D059467:Transcriptome; D014733:Vibrio / Q000502:physiology; D046848:White spot syndrome virus 1 / Q000502:physiology
chemical_list: D060829:Arthropod Proteins; D018076:DNA, Complementary; D017346:Protein Serine-Threonine Kinases