title: Sulfated galactans from Gracilaria fisheri bind to shrimp haemocyte membrane proteins and stimulate the expression of immune genes.
abstract: Previous studies demonstrated that sulfated galactans (SG) from Gracilaria fisheri (G. fisheri) exhibit immunostimulant activity in shrimp. The present study was conducted to test the hypothesis that SG stimulates signaling molecules of the immune response of shrimp by binding to receptors on the host cell membrane. Accordingly, we evaluated the ability of SG to bind to shrimp haemocytes and showed that SG bound to the shrimp haemocyte membrane (SHM), potentially to specific receptors. Furthermore, this binding was associated with an activation of immune response genes of shrimp. Data from confocal laser scanning micrographs revealed that FITC-labeled SG bound to haemocytes. Far western blot analysis demonstrated that SHM peptides, with molecular sizes of 13, 14, 15, 17, and 25 kDa, were associated with SG. Peptide sequence analysis of the isolated bands using LC-MS/MS and NCBI blast search revealed the identity of the 13, 14, and 17 kDa peptides as lipopolysaccharide and β-1,3-glucan binding protein (LGBP). SG induced the expression of immune related genes and downstream signaling mediators of LGBP including IMD, IKKs, NF-κB, antimicrobial peptides (crustin and PEN-4), the antiviral immunity (dicer), and proPO system (proPO-I and proPO-II). A LGBP neutralizing assay with anti-LGBP antibody indicated a decrease in SG-induced expression of LGBP downstream signaling mediators and the immune related genes. In conclusion, this study demonstrated that the SG-stimulated immune activity in haemocytes is mediated, in part, through the LGBP, and IMD-NF-κB pathway.
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama 6th Road, Bangkok 10400, Thailand.
authors.forename: Tawut
authors.initials: T
authors.lastname: Rudtanatip
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama 6th Road, Bangkok 10400, Thailand; Centex Shrimp, Faculty of Science, Mahidol University, Rama 6th Road, Bangkok 10400, Thailand; The Shrimp Genetic Improvement Center, Chaiya District, Surat Thani 84100, Thailand.
authors.forename: Boonsirm
authors.initials: B
authors.lastname: Withyachumnarnkul
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama 6th Road, Bangkok 10400, Thailand. Electronic address: kanokpan.won@mahidol.ac.th.
authors.forename: Kanokpan
authors.initials: K
authors.lastname: Wongprasert
mesh_terms: D000818:Animals; D060829:Arthropod Proteins / Q000235:genetics* / Q000378:metabolism; D005685:Galactans / Q000737:chemistry / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D044724:Gracilaria / Q000737:chemistry*; D006434:Hemocytes / Q000187:drug effects / Q000378:metabolism; D007113:Immunity, Innate; D008565:Membrane Proteins / Q000378:metabolism*; D033561:Penaeidae / Q000187:drug effects* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013455:Sulfur / Q000737:chemistry
chemical_list: D060829:Arthropod Proteins; D005685:Galactans; D008565:Membrane Proteins; D013455:Sulfur
keywords: Gracilaria fisheri; Haemocyte culture; Immune response; LGBP; Shrimp haemocyte membrane proteins; Sulfated galactans