Transcriptional Evidence for the Involvement of MyD88 in Flagellin Recognition: Genomic Identification of Rock Bream MyD88 and Comparative Analysis
The MyD88 is an evolutionarily conserved host-expressed adaptor protein that is essential for proper TLR/ IL1R immune-response signaling. A previously identified complete cDNA (1626 bp) of OfMyD88 comprised an ORF of 867 bp encoding a protein of 288 amino acids (32.9 kDa). The gDNA (3761 bp) of OfMyD88 revealed a quinquepartite genome organization composed of 5 exons (with the sizes of 310, 132, 178, 92 and 155 bp) separated by 4 introns. All the introns displayed splice signals consistent with the consensus GT/AG rule. A bipartite domain structure with two domains namely death domain (24-103) coded by 1st exon, and TIR domain (151-288) coded by last 3 exons were identified through in silico analysis. Moreover, homology modeling of these two domains revealed a similar quaternary folding nature between human and rock bream homologs. A comprehensive comparison of vertebrate MyD88 genes showed that they possess a 5-exonic structure.In this structure, the last three exons were strongly conserved, and this suggests that a rigid structure has been maintained during vertebrate evolution.A cluster of TATA box-like sequences were found 0.25 kb upstream of cDNA starting position. In addition, putative 5'-flanking region of OfMyD88 was predicted to have TFBS implicated with TLR signaling, including copies of NFkB1, APRF/ STAT3, Sp1, IRF1 and 2 and Stat1/2. Using qPCR technique, a ubiquitous mRNA expression was detected in liver and blood. Furthermore, a significantly up-regulated transcriptional expression of OfMyD88 was detected in head kidney (12-24 h; >2-fold), spleen (6 h; 1.5-fold), liver (3 h; 1.9-fold) and intestine (24 h; ~2-fold) post-Fla challenge. These data suggest a crucial role for MyD88 in antibacterial immunity of teleosts.
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 Armant MA, Fenton MJ. "Toll-like receptors: a family of pattern-recognition receptors in mammals." Genome biology, vol. 3, 2002, pp. reviews3011.
 Beutler B. "The Toll-like receptors: analysis by forward genetic methods." Immunogenetics, vol. 57, 2005, pp. 385-92.
 Bilak H, Tauszig-Delamasure S, Imler JL. "Toll and Toll-like receptors in Drosophila."BiochemSoc Trans, vol. 31, 2003, pp. 648-51.
 Krishnan J, Selvarajoo K, Tsuchiya M, Lee G, Choi S. "Toll-like receptor signal transduction." ExpMol Med, vol. 39, 2007, pp. 421-38.
 Janssens S, Beyaert R. "A universal role for MyD88 in TLR/IL-1R-mediated signaling." Trends in biochemical sciences, vol. 27, 2002, pp. 474-82.
 Medzhitov R, Preston-Hurlburt P, Kopp E, Stadlen A, Chen C, Ghosh S, et al. "MyD88 is an adaptor protein in the hToll/IL-1 receptor family signaling pathways." Molecular cell, vol. 2, 1998, pp. 253-8.
 Wesche H, Henzel WJ, Shillinglaw W, Li S, Cao Z. "MyD88: an adapter that recruits IRAK to the IL-1 receptor complex." Immunity, vol. 7, 1997, pp. 837-47.
 van der Sar AM, Stockhammer OW, van der Laan C, Spaink HP, Bitter W, Meijer AH. "MyD88 innate immune function in a zebrafish embryo infection model."Infection and immunity, vol. 74, 2006, pp. 2436-41.
 Takano T, Kondo H, Hirono I, Saito-Taki T, Endo M, Aoki T. "Identification and characterization of a myeloid differentiation factor 88 (MyD88) cDNA and gene in Japanese flounder, Paralichthysolivaceus." Developmental & Comparative Immunology, vol. 30, 2006, pp. 807-16.
 Yao CL, Kong P, Wang ZY, Ji PF, Liu XD, Cai MY, et al. "Molecular cloning and expression of MyD88 in large yellow croaker, Pseudosciaenacrocea." Fish Shellfish Immunol, vol. 26, 2009, pp. 249-55.
 Rebl A, Goldammer T, Fischer U, Kollner B, Seyfert HM. "Characterization of two key molecules of teleost innate immunity from rainbow trout (Oncorhynchusmykiss): MyD88 and SAA." Vet ImmunolImmunopathol, vol. 131, 2009, pp. 122-6.
 Whang I, Lee Y, Kim H, Jung SJ, Oh MJ, Choi CY, et al. "Characterization and expression analysis of the myeloid differentiation factor 88 (MyD88) in rock bream Oplegnathusfasciatus." MolBiol Rep, vol., 2010, pp. 3911-3920.
 Umasuthan N, Whang I, Kim J-O, Oh M-J, Jung S-J, Choi CY, et al. "Rock bream (Oplegnathusfasciatus) serpin, protease nexin-1: Transcriptional analysis and characterization of its antiprotease and anticoagulant activities." Developmental & Comparative Immunology, vol. 35, 2011, pp. 785-98.
 Umasuthan N, Bathige S, Revathy KS, NiroshanaWickramaarachchi W, Wan Q, Whang I, et al. "AC1 inhibitorortholog from rock bream (Oplegnathusfasciatus): Molecular perspectives of a central regulator in terms of its genomic arrangement, transcriptional profiles and anti-protease activities of recombinant peptide." Developmental & Comparative Immunology, vol., 2013, pp. 197-210.
 Lynn DJ, Lloyd AT, O’Farrelly C. "In silicoidentification of components of the Toll-like receptor (TLR) signaling pathway in clustered chicken expressed sequence tags (ESTs)." Veterinary Immunology and Immunopathology, vol. 93, 2003, pp. 177-84.
 Prothmann C, Armstrong NJ, Rupp RA. "The Toll/IL-1 receptor binding protein MyD88 is required for Xenopus axis formation." MechDev, vol. 97, 2000, pp. 85-92.
 Harroch S, Gothelf Y, Revel M, Chebath J. "5′ upstream sequences of MyD88, an IL-6 primary response gene in M1 cells: detection of functional IRF-1 and Stat factors binding sites." Nucleic acids research, vol. 23, 1995, pp. 3539-46.
 Basu M, Swain B, Maiti NK, Routray P, Samanta M. "Inductive expression of toll-like receptor 5 (TLR5) and associated downstream signaling molecules following ligand exposure and bacterial infection in the Indian major carp, mrigal (Cirrhinusmrigala)." Fish & Shellfish Immunology, vol. 32, 2012, pp. 121-31.