Abstract：Heart-type fatty acid-binding protein (FABP-3) is a member of the lipid-binding protein superfamily, and its main functions are uptaking and transportation of fatty acids. In order to understand the structure of FABP-3 and its potential function in nutrition metabolism and immune regulation of Acipenser dabryanus, the full-length cDNA of A. dabryanus FABP-3 was cloned from the unigenes database by full-length transcriptome sequencing of A. dabryanus. The expression profiles of FABP-3 in different tissues, starvation stress and bacterial infection were analyzed. The full length of FABP-3 was 641 bp (GenBank No. MN064725) which contained 89 bp 5' untranslated region (UTR), 150 bp 3' UTR and 402 bp open reading frame (ORF). The ORF of FABP-3 encoded 133-residue peptide. FABP-3 was predicted to have a lipocalin domain from the 6 to 120 amino acids by analysis of the deduced protein sequence using the SMART program. The results also showed the three-dimensional protein structure model predicted by SWISS-MODEL contained 10 antiparallel β-strands and 2 α-helixes in the lipocalin domain, and the 3 conserved amino acid residues (Arg 107, Arg 127, Tyr129) were also present. The tissue distribution and expression level during starvation stress were further analyzed. Results showed that FABP-3 was ubiquitously expressed in all tested tissues, and was mostly expressed in muscle (P≤0.05). During starvation, the expression level of FABP-3 in the brain, liver and stomach increased first and then decreased, while the expression of FABP-3 in the intestine and muscle declined significantly compared to the control group (P≤0.05). Bacterial challenge in vivo revealed significant changes in the gene expression of FABP-3 at immune-related sites. Striking upregulation of FABP-3 was observed in the hindgut at 12 h post-challenge, with increases by more than 219-fold compared to the control, and the expression of FABP-3 mRNA was decreased at first and remarkably elevated in the gill and spleen at 48 h post-challenge. The results indicated that FABP-3 was involved in biological pathways such as nutrition metabolism and immune response in A. dabryanus and would provide basic information for future studies on nutritional and immunomodulatory molecular mechanisms of the A. dabryanus.
陈叶雨, 刘亚, 赖见生, 宋明江, 龚全. 达氏鲟心型脂肪酸结合蛋白的基因克隆及表达分析[J]. 农业生物技术学报, 2020, 28(1): 139-149.
CHEN Ye-Yu, LIU Ya, LAI Jian-Sheng, SONG Ming-Jiang, GONG Quan. Gene Cloning and Expression Analysis of Heart-type Fatty Acid-binding Protein in Acipenser dabryanus. 农业生物技术学报, 2020, 28(1): 139-149.
[1] 高红, 彭静, 沈一飞, 等. 2014. 不同物种FABP3基因编码区及蛋白生物信息学分析[J]. 扬州大学学报, 35(1): 31-37. (Gao H, Peng J, Shen Y F, et al.2014. Bioinformatic analysis of coding regions and proteins of FABP3 gene among different species[J]. Journal of Yangzhou University, 35(1): 31-37.) [2] 高露姣, 陈立侨, 宋兵. 2004. 饥饿和补偿生长对史氏鲟幼鱼摄食、生长和体成分的影响[J]. 水产学报, 28(3): 279-284. (Gao L J, Chen L Q, Song B.2004. Effect of starvation and compensatory growth on feeding, growth and body biochemical composition in Acipenser schrenckii juveniles[J]. Journal of Fisheries of China, 28(3): 279-284.) [3] 黄玉海, 汪以真, 单体中, 等. 2006. 猪心型脂肪酸结合蛋白基因的克隆及不同生长阶段表达的差异[J]. 农业生物技术学报, 14(5): 657-661. (Huang Y H, Wang Y Z, Shan T Z, et al.2006. Cloning of heart-type fatty acid-binding protein (H-FABP) gene of swine and difference of its expression at different stages[J]. Journal of Agricultural Biotechnology, 14(5): 657-661.) [4] 姜志强, 贾泽梅, 韩延波. 2002. 美国红鱼继饥饿后的补偿生长及其机制[J]. 水产学报, 26(1): 67-72. (Jiang Z Q, Jia Z M, Han Y B.2002. The compensatory growth and its mechanismin of red drum, Sciaenops ocellatus, after food deprivation[J]. Journal of Fisheries of China, 26(1): 67-72.) [5] 梁梦瑶, 高学军, 李庆章, 等. 2013. 脂肪酸结合蛋白3在乳脂合成信号转导通路中的研究进展[J]. 中国畜牧兽医, 40(12): 116-120. (Liang M Y, Gao X J, Li Q Z, et al.2013. Research progress on the milk fat synthesis signal transduction pathway of FABP3[J]. China Animal Husbandry and Veterinary Medicine, 40(12): 116-120.) [6] 廖玉英, 叶超霞, 尹嘉琪, 等. 2014. 暗纹东方鲀心型脂肪酸结合蛋白H-FABP基因克隆与表达分析[J]. 水产学报, 11(38): 1808-1818. (Liao Y Y, Ye C X, Yin J Q, et al.2014. Molecular cloning and feature analysis of heart-type fatty acid-binding protein H-FABP gene from obscure puffer (Takifugu obscurus)[J]. Journal of Fisheries of China, 11(38): 1808-1818.) [7] 林亚秋, 李瑞文, 郑玉才, 等. 2011. 齐口裂腹鱼和鲤鱼H-FABP基因的克隆及其表达谱[J]. 中国生物化学与分子生物学报, 27(6): 554-560. (Lin Y Q, Li R W, Zheng Y C, et al.2011. Cloning and expression profile of heart-type fatty acid-binding protein in Schizothorax prenanti and Cyprinus carpio[J]. Chinese Journal of Biochemistry and Molecular Biology, 27(6): 554-560.) [8] 刘波, 何庆国, 唐永凯, 等. 2009. 饥饿胁迫对吉富罗非鱼生长及生理生化指标的影响[J]. 中国水产科学, 16(2): 230-237. (Liu B, He Q G, Tang Y K, et al.2009. Effects of starvation on growth, physiological and biochemical parameter of GIFT strain of Nile tilapia (Oreochromis niloticus)[J]. Journal of Fishery Sciences of China, 16(2): 230-237.) [9] 刘晓勇, 张颖, 齐茜, 等. 2011. 鲟鱼营养与饲料的研究进展和发展趋势[J]. 饲料工业, 32(20): 25-28. (Liu X Y, Zhang Y, Qi Q, et al.2011. Research progress and development trend of nutrition and feed of sturgeon[J]. Feed Industry, 32(20): 25-28.) [10] 刘亚, 龚全, 李强, 等. 2017. 全人工繁殖达氏鲟胚胎发育的形态学和组织学观察[J]. 西南农业学报, 30(7): 1686-1692. (Liu Y, Gong Q, Li Q, et al.2017. Morphological and histological observation of embryonic development of artificially-bred Acipenser dabryanus Dumeril[J]. Southwest China Journal of Agricultural Sciences, 30(7): 1686-1692.) [11] 覃川杰, 邵婷, 杨洁萍, 等. 2015. 饥饿胁迫对瓦氏黄颡鱼脂肪代谢的影响[J]. 水生生物学报, 39(1): 58-65. (Qin C J, Shao T, Yang J P, et al.2015. The effect of starvation on lipid metabolism of darkbarbel catfish, Pelteobagrus vachelli[J]. Acta Hydrobiologica Sinica, 39(1): 58-65.) [12] 谢小军, 邓利, 张波. 1998. 饥饿对鱼类生理生态学影响的研究进展[J]. 水生生物学报, 22(2): 181-189. (Xie X J, Deng L, Zhang B.1998. Advances and studies on ecophysiological effects of starvation on fish[J]. Acta Hydrobiologica Sinica, 22(2): 181-189.) [13] 赵永贞, 陈秀荔, 谢达祥, 等. 2014. 凡纳滨对虾脂肪酸结合蛋白基因全长cDNA的克隆及序列分析[J]. 水产学报, 34(11): 1681-1688. (Zhao Y Z, Chen X L, Xie D X, et al.2014. Cloning and analysis of full length cDNA of intracellular fatty-acid-binding protein in Litopenaeus vannamei[J]. Journal of Fisheries of China, 34(11): 1681-1688.) [14] Ando S, Xue X H, Tibbits G F, et al.1998. Cloning and sequencing of complementary DNA for fatty acid binding protein from rainbow trout heart[J]. Comparative Biochemistry & Physiology Part B: Biochemistry & Molecular Biology, 119(1): 213-217. [15] Bain L J.2002. cDNA cloning, sequencing, and differential expression of a heart-type fatty acid-binding protein in the mummichog (Fundulus heteroclitus)[J]. Marine Environmental Research, 54(3): 379-383. [16] Bayır M, Bayır A, Wright J M.2015. Divergent spatial regulation of duplicated fatty acid-binding protein (fabp) genes in rainbow trout (Oncorhynchus mykiss)[J]. Comparative Biochemistry and Physiology Part D: Genomics and Proteomics, 14: 26-32. [17] Chen W B, Wang K M, Liu S Y.2018. Molecular cloning and tissue distribution of fatty acid binding protein-3 in goldfish (Carassius auratus) and its mRNA expression in response to cadmium and PAMPs[J]. Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 224: 68-75. [18] Chen Y Y, Gong Q, Song M J, et al.2019. Identification and characterization of three novel antimicrobial peptides from Acipenser dabryanus[J]. Fish & Shellfish Immunology, 88: 207-216. [19] Esteves A, Ehrlich R.2006. Invertebrate intracellular fatty acid binding proteins[J]. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology, 142(3): 262-274. [20] Gerbens F, Jansen A, Erp A J M V, et al.1998. The adipocyte fatty acid-binding protein locus: characterization and association with intramuscular fat content in pigs[J]. Mammalian Genome, 9(12): 1022-1026. [21] Hittel D, Storey K B.2001. Differential expression of adipose- and heart-type fatty acid binding proteins in hibernating ground squirrels[J]. Biochimica et Biophysica Acta, 1522(3): 238-243. [22] Hung S, Liu W, Li H, et al.1997. Effect of starvation on some morphological and biochemical parameters in white sturgeon, Acipenser transmontanus[J]. Aquaculture, 151: 357-363. [23] Liu R Z, Saxena V, Sharma M K, et al.2007. The fabp4 gene of zebrafish (Danio rerio)--genomic homology with the mammalian FABP4 and divergence from the zebrafish fabp3 in developmental expression[J]. FEBS Journal, 274(6): 1621-1633. [24] Liu R Z, Wright D, Wright J M.2003. Structure, linkage mapping and expression of the heart-type fatty acid-binding protein gene (fabp3) from zebrafish (Danio rerio)[J]. European Journal of Biochemistry, 270: 3223-3224. [25] Parmar M B, Wright J M.2013. Comparative genomic organization and tissue-specific transcription of the duplicated fabp7 and fabp10 genes in teleost fishes[J]. Genome, 56(11): 691-701. [26] Selsted M E, Tang Y Q, Morris W L, et al.1996. Purification, primary structures, and antibacterial activities of beta-defensins, a new family of antimicrobial peptides from bovine neutrophils[J]. The Journal of Biological Chemistry, 271: 16430. [27] Tan S J, Zhang X, Jin X K, et al.2015. Fatty acid binding protein FABP3 from Chinese mitten crab Eriocheir sinensis participates in antimicrobial responses[J]. Fish & Shellfish Immunology, 43(1): 264-274. [28] Venkatachalam A B, Fontenot Q, Farrara A, et al.2018. Fatty acid-binding protein genes of the ancient, air-breathing, ray-finned fish, spotted gar (Lepisosteus oculatus)[J]. Comparative Biochemistry and Physiology Part D: Genomics & Proteomics, 25: 19-25. [29] Venkatachalam A B, Sawler D L, Wright J M.2013. Tissue-specific transcriptional modulation of fatty acid-binding protein genes, fabp2, fabp3 and fabp6, by fatty acids and the peroxisome proliferator, clofibrate, in zebrafish (Danio rerio)[J]. Gene, 520(1): 14-21. [30] Wang S, Zhu Y, Li X, et al.2017. Fatty acid binding protein regulate antimicrobial function via Toll signaling in Chinese mitten crab[J]. Fish & Shellfish Immunology, 63: 9-17. [31] Wu J M, Wei Q W, Du H, et al.2015. Initial evaluation of the release programme for Dabry's sturgeon (Acipenser dabryanus Duméril, 1868) in the upper Yangtze River[J]. Journal of Applied Ichthyology, 30(6): 1423-1427. [32] Yang R, Liu Y, Wang Y, et al.2018. Pathogenesis and pathological analysis of Edwardsiella tarda from Dabry's sturgeon (Acipenser dabryanus) in China[J]. Aquaculture, 495: 637-642. [33] Yount N Y, Yuan J, Tarver A, et al.1999. Cloning and expression of bovine neutrophil beta-defensins. Biosynthetic profile during neutrophilic maturation and localization of mature peptide to novel cytoplasmic dense granules[J]. The Journal of Biological Chemistry, 274: 26249-26258.
