title: Cloning and characterisation of the SpToll gene from green mud crab, Scylla paramamosain.
abstract: Toll/Toll-like receptors (TLRs), one of the most important pattern recognition receptors (PRRs), play a crucial role in innate immune responses in both invertebrates and vertebrates. In this study, we cloned and characterised a Toll gene from Scylla paramamosain (SpToll). Bioinformatic analysis predicted that SpToll contained one open reading frame of 3018bp and encoded a single-pass transmembrane domain protein of 1005 amino acids. Further, SpToll could be clustered into one branch along with other arthropod Tolls in a phylogenetic tree. SpToll transcripts could be detected by RT-PCR from all tissues examined including the heart, gill, hepatopancreas, stomach, intestine, muscle, eyestalk and hemocytes. Infection by Vibrio parahemolyticus up-regulated SpToll mRNA expression in hemocytes after 48h. The profile of single nucleotide polymorphisms (SNPs) in the leucine-rich repeats (LRRs) domain of SpToll in three healthy crabs was then evaluated. Two hundred and twenty SNPs with a frequency of about 1.0-4.0% were identified in hemocyte DNA/cDNA. Surprisingly, the adenine to guanine transition at position 1372 (c.1372A>G) had a frequency of about 50%. Finally, the results showed that challenge with V. parahemolyticus stimulated the appearance of two sets of SNPs in crabs. More importantly, the c.1372A>G mutation could contribute to a low mortality after V. parahemolyticus infection and introduce variation of charge and secondary structure into the SpToll polypeptide. In summary, these studies suggested a novel Toll homologue in crab and identified a SNP with potential pathogen-resistant activities. The result will be important for the investigation of crab immune defense mechanisms.
authors.affiliation: Department of Biology and Guangdong Provincial Key Laboratory of Marine Biotechnology, Shantou University, Shantou, Guangdong 515063, China.
authors.forename: Zhongyang
authors.initials: Z
authors.lastname: Lin
authors.forename: Jie
authors.initials: J
authors.lastname: Qiao
authors.forename: Yueling
authors.initials: Y
authors.lastname: Zhang
authors.forename: Lingling
authors.initials: L
authors.lastname: Guo
authors.forename: He
authors.initials: H
authors.lastname: Huang
authors.forename: Fang
authors.initials: F
authors.lastname: Yan
authors.forename: Yuanyou
authors.initials: Y
authors.lastname: Li
authors.forename: Xiuying
authors.initials: X
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060829:Arthropod Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003386:Brachyura / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D016384:Consensus Sequence; D060467:Disease Resistance / Q000235:genetics; D005786:Gene Expression Regulation; D006434:Hemocytes / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D020641:Polymorphism, Single Nucleotide; D017434:Protein Structure, Tertiary; D017422:Sequence Analysis, DNA; D051193:Toll-Like Receptors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D014735:Vibrio Infections / Q000276:immunology / Q000378:metabolism / Q000662:veterinary
chemical_list: D060829:Arthropod Proteins; D051193:Toll-Like Receptors