• ISSN 1000-0615
  • CN 31-1283/S
Volume 47 Issue 8
Aug.  2023
Turn off MathJax
Article Contents
Abstract
References
WANG Yujing, XU Yuxin, FENG Wenrong, LI Jianlin, LI Hongxia, SU Shengyan, SONG Changyou, TANG Yongkai, YU Juhua. Sequence analysis, expression patterns, acquisition of prokaryotic recombinant protein and pro-inflammatory effect of IL-17B in Cyprinus carpio[J]. Journal of fisheries of china, 2023, 47(8): 089103. DOI: 10.11964/jfc.20210812990
Citation: WANG Yujing, XU Yuxin, FENG Wenrong, LI Jianlin, LI Hongxia, SU Shengyan, SONG Changyou, TANG Yongkai, YU Juhua. Sequence analysis, expression patterns, acquisition of prokaryotic recombinant protein and pro-inflammatory effect of IL-17B in Cyprinus carpio[J]. Journal of fisheries of china, 2023, 47(8): 089103. DOI: 10.11964/jfc.20210812990
shu

Sequence analysis, expression patterns, acquisition of prokaryotic recombinant protein and pro-inflammatory effect of IL-17B in Cyprinus carpio

  • 1.

    Wuxi Fisheries College, Nanjing Agricultural University, Wuxi　214128, China

  • 2.

    Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi　214081, China

Funds: Central Public-interest Scientific Institution Basal Research Fund, CAFS (2018HY-ZD02, 2020TD37); China Agriculture Research System of MOF and MARA(CARS-45)
More Information
  • Corresponding author:

    TANG Yongkai. E-mail: tangyk@ffrc.cn;

    YU Juhua. E-mail: yujh@ffrc.cn

  • Received Date: August 04, 2021
  • Revised Date: September 08, 2021
  • Available Online: July 14, 2023
  • Published Date: July 31, 2023
    Graphical Abstract
    • Abstract
  • Interleukin-17 (IL-17) is an important inflammatory factor, which is able to promote the production of various inflammation-related factors, such as IL-1β, IFN-γ, IL-6, CCL20 and so forth. It plays an important role in autoimmune disease, host defense, inflammation and so on. In order to study the pro-inflammatory function of IL-17B in the Cyprinus carpio, the IL-17B gene was cloned, the NusA-17B protein was recombinantly expressed by Escherichia coli system and the pro-inflammatory effect of the IL-17B was then investigated by in vivo and in vitro experiments. In this study, by homology search and gene cloning method, we found two IL-17Bs in the genome of the C. carpio, CcIL-17B1 and CcIL-17B2 respectively. Both of the two genes have open reading frames (ORF) of 597 bp, which are composed of three exons and two introns. The length of their exons are the same, 24, 320 and 253 bp respectively. While the introns are different, the CcIL-17B1 intron are 84 , 420 bp respectively and the CcIL-17B2 intron are 81, 851 bp respectively. The two CcIL-17Bs encode 198 amino acids, containing 2 disulfide bonds formed by 4 cysteines, which were unique to the IL-17 family. Sequence blast analysis showed that the consistency of the two CcIL-17Bs proteins reached 91.92%. The collinearity analysis results are as follows. During the doubling of the chromosomes of teleost, IL-17B and its nearby genes were lost. Most teleost have one IL-17B and Danio rerio have both IL-17Bs that were lost. The common carp-specific chromosomes doubled resulting in the presence of two CcIL-17Bs. Real time quantitative PCR (qPCR) was used to measure the expression of CcIL-17B1 and CcIL-17B2 in fertilized eggs, larvae and different tissues of adult C. carpio. The results showed that the expression levels of both CcIL-17B1 and CcIL-17B2 were significantly higher at 0-12 hours post fertilization than that at 25-90 hours post fertilization (P<0.05). While the expression levels have no significant difference at 25-90 hours after fertilization and 1-14 days after hatching (P>0.05). In adult fish, the expression level of two genes in testis and gonad are significantly higher than that in other tissues, such as spleen, heart, kidney, head kidney, intestine, gill, liver, skin, muscle, brain (P<0.05). Further, using the E. coli expression system, the soluble recombinant protein NusA-17B was obtained. Different concentrations of NusA-17B were injected into the anus to evaluate its pro-inflammatory effect. Histopathological results showed that intestinal villi defect, plenty of goblet cells and inflammatory cells appeared in intestines treated with high concentration NusA-17B (500 μg/kg) after 1 and 3 days. Meanwhile, qPCR showed that IL-1β, IFN-γ, IL-6, CCL20 and NF-κB were significantly up-regulated (P<0.05). However, the abnormal changes in intestinal structure and expression of inflammation-related genes were restored to normal level after 7 days. In addition, in vitro, the C. carpio kidney cells were incubated into different concentrations (0.1, 1.0, 10.0 and 100.0 ng/mL) NusA-17B for 8 hours. The results showed that IL-1β was significantly up-regulated under NusA-17B stimulation at 1.0, 10. and 100.0 ng/mL (P<0.05); IFN-γ and NF-κB were significantly up-regulated under the stimulation of NusA-17B at 10.0 ng/mL and 0.1 ng/mL respectively (P<0.05); IL-6 and CCL20 were significantly expressed under NusA-17B stimulation at 10.0 and 100.0 ng/mL (P<0.05), and TRAF6 was significantly expressed under NusA-17B stimulation at 0.1, 1.0 and 10.0 ng/mL (P<0.05). In summary, the results of in vivo and in vitro experiments show that CcIL-17B participates in the inflammatory response.
    • FullText(HTML)
    • References (45)
  • [1]
    Miossec P, Korn T, Kuchroo V K. Interleukin-17 and type 17 helper T cells[J]. New England Journal of Medicine, 2009, 361(9): 888-898. doi: 10.1056/NEJMra0707449
    [2]
    Kawaguchi M, Adachi M, Oda N, et al. IL-17 cytokine family[J]. Journal of Allergy and Clinical Immunology, 2004, 114(6): 1265-1273. doi: 10.1016/j.jaci.2004.10.019
    [3]
    Rouvier E, Luciani M F, Mattéi M G, et al. CTLA-8, cloned from an activated T cell, bearing AU-rich messenger RNA instability sequences, and homologous to a herpesvirus saimiri gene[J]. Journal of Immunology, 1993, 150(12): 5445-5456. doi: 10.4049/jimmunol.150.12.5445
    [4]
    Gerhardt S, Abbott W M, Hargreaves D, et al. Structure of IL-17A in complex with a potent, fully human neutralizing antibody[J]. Journal of Molecular Biology, 2009, 394(5): 905-921. doi: 10.1016/j.jmb.2009.10.008
    [5]
    Hymowitz S G, Filvaroff E H, Yin J P, et al. IL-17s adopt a cystine knot fold: structure and activity of a novel cytokine, IL-17F, and implications for receptor binding[J]. The EMBO Journal, 2001, 20(19): 5332-5341. doi: 10.1093/emboj/20.19.5332
    [6]
    Gunimaladevi I, Savan R, Sakai M. Identification, cloning and characterization of interleukin-17 and its family from zebrafish[J]. Fish and Shellfish Immunology, 2006, 21(4): 393-403. doi: 10.1016/j.fsi.2006.01.004
    [7]
    Kumari J, Larsen A N, Bogwald J, et al. Interleukin-17D in Atlantic salmon (Salmo salar): molecular characterization, 3D modelling and promoter analysis[J]. Fish and Shellfish Immunology, 2009, 27(5): 647-659. doi: 10.1016/j.fsi.2009.08.009
    [8]
    Korenaga H, Kono T, Sakai M. Isolation of seven IL-17 family genes from the Japanese pufferfish Takifugu rubripes[J]. Fish and Shellfish Immunology, 2010, 28(5-6): 809-818. doi: 10.1016/j.fsi.2010.01.016
    [9]
    Kono T, Korenaga H, Sakai M. Genomics of fish IL-17 ligand and receptors: a review[J]. Fish and Shellfish Immunology, 2011, 31(5): 635-643. doi: 10.1016/j.fsi.2010.11.028
    [10]
    曹永生, 徐黎明, 赵景壮, 等. 虹鳟IL-17多克隆抗体的制备及初步鉴定[J]. 水产学杂志, 2016, 29(6): 1-5. doi: 10.3969/j.issn.1005-3832.2016.06.001

    Cao Y S, Xu L M, Zhao J Z, et al. Preparation and identification of polyclonal antibody of IL-17 in rainbow trout (Oncorhynchus mykiss)[J]. Chinese Journal of Fisheries, 2016, 29(6): 1-5 (in Chinese). doi: 10.3969/j.issn.1005-3832.2016.06.001
    [11]
    Monte M M, Wang T H, Holland J W, et al. Cloning and characterization of rainbow trout interleukin-17A/F2 (IL-17A/F2) and IL-17 receptor A: expression during infection and bioactivity of recombinant IL-17A/F2[J]. Infection and Immunity, 2013, 81(1): 340-353. doi: 10.1128/IAI.00599-12
    [12]
    Wang X Q, Li C, Thongda W, et al. Characterization and mucosal responses of interleukin 17 family ligand and receptor genes in channel catfish Ictalurus punctatus[J]. Fish and Shellfish Immunology, 2014, 38(1): 47-55. doi: 10.1016/j.fsi.2014.02.020
    [13]
    Nuñez Ortiz N, Gerdol M, Stocchi V, et al. T cell transcripts and T cell activities in the gills of the teleost fish sea bass (Dicentrarchus labrax)[J]. Developmental and Comparative Immunology, 2014, 47(2): 309-318. doi: 10.1016/j.dci.2014.07.015
    [14]
    Du L Y, Feng S Y, Yin L C, et al. Identification and functional characterization of grass carp IL-17A/F1: an evaluation of the immunoregulatory role of teleost IL-17A/F1[J]. Developmental and Comparative Immunology, 2015, 51(1): 202-211. doi: 10.1016/j.dci.2015.03.014
    [15]
    Du L Y, Qin L, Wang X Y, et al. Characterization of grass carp (Ctenopharyngodon idella) IL-17D: molecular cloning, functional implication and signal transduction[J]. Developmental and Comparative Immunology, 2014, 42(2): 220-228. doi: 10.1016/j.dci.2013.09.015
    [16]
    Yang Q, Sun Y N, Su X R, et al. Characterization of six IL-17 family genes in miiuy croaker and evolution analysis of vertebrate IL-17 family[J]. Fish and Shellfish Immunology, 2016, 49: 243-251. doi: 10.1016/j.fsi.2015.12.031
    [17]
    Dong C J, Kong S N, Zheng X H, et al. Genome-wide identification of interleukin-17 (IL17) in common carp (Cyprinus carpio) and its expression following Aeromonas hydrophila infection[J]. Gene, 2019, 686: 68-75. doi: 10.1016/j.gene.2018.10.038
    [18]
    Li H X, Yu J H, Li J L, et al. Cloning and characterization of two duplicated interleukin-17A/F2 genes in common carp (Cyprinus carpio L. ): transcripts expression and bioactivity of recombinant IL-17A/F2[J]. Fish and Shellfish Immunology, 2016, 51: 303-312. doi: 10.1016/j.fsi.2016.01.042
    [19]
    Li H Z, Chen J, Huang A, et al. Cloning and characterization of IL-17B and IL-17C, two new members of the IL-17 cytokine family[J]. Proceedings of the National Academy of Sciences of the United States of America, 2000, 97(2): 773-778. doi: 10.1073/pnas.97.2.773
    [20]
    Shi Y G, Ullrich S J, Zhang J, et al. A novel cytokine receptor-ligand pair: identification, molecular characterization, and in vivo immunomodulatory activity[J]. Journal of Biological Chemistry, 2000, 275(25): 19167-19176. doi: 10.1074/jbc.M910228199
    [21]
    Iwakura Y, Ishigame H, Saijo S, et al. Functional specialization of interleukin-17 family members[J]. Immunity, 2011, 34(2): 149-162. doi: 10.1016/j.immuni.2011.02.012
    [22]
    Kolls J K, Lindén A. Interleukin-17 family members and inflammation[J]. Immunity, 2004, 21(4): 467-476. doi: 10.1016/j.immuni.2004.08.018
    [23]
    Gaffen S L. Structure and signalling in the IL-17 receptor family[J]. Nature Reviews Immunology, 2009, 9(8): 556-567. doi: 10.1038/nri2586
    [24]
    Kouri V P, Olkkonen J, Ainola M, et al. Neutrophils produce interleukin-17B in rheumatoid synovial tissue[J]. Rheumatology, 2014, 53(1): 39-47. doi: 10.1093/rheumatology/ket309
    [25]
    Moore E E, Presnell S, Garrigues U, et al. Expression of IL-17B in neurons and evaluation of its possible role in the chromosome 5q-linked form of Charcot-Marie-Tooth disease[J]. Neuromuscular Disorders, 2002, 12(2): 141-150. doi: 10.1016/S0960-8966(01)00250-4
    [26]
    Kokubu T, Haudenschild D R, Moseley T A, et al. Immunolocalization of IL-17A, IL-17B, and their receptors in chondrocytes during fracture healing[J]. Journal of Histochemistry and Cytochemistry, 2008, 56(2): 89-95. doi: 10.1369/jhc.7A7223.2007
    [27]
    Ryan A W, Thornton J M, Brophy K, et al. Chromosome 5q candidate genes in coeliac disease: genetic variation at IL4, IL5, IL9, IL13, IL17B and NR3C1[J]. Tissue Antigens, 2005, 65(2): 150-155. doi: 10.1111/j.1399-0039.2005.00354.x
    [28]
    Li H X, Zhang L, Li J L, et al. Identification, expression and pro-inflammatory effect of interleukin-17 N in common carp (Cyprinus carpio L. )[J]. Fish and Shellfish Immunology, 2021, 111: 6-15. doi: 10.1016/j.fsi.2020.11.024
    [29]
    Dong X B, Tang B, Li J, et al. Expression and purification of intact and functional soybean (Glycine max) seed ferritin complex in Escherichia coli[J]. Journal of Microbiology and Biotechnology, 2008, 18(2): 299-307.
    [30]
    Young C L, Britton Z T, Robinson A S. Recombinant protein expression and purification: a comprehensive review of affinity tags and microbial applications[J]. Biotechnology Journal, 2012, 7(5): 620-634. doi: 10.1002/biot.201100155
    [31]
    Song X H, Zhao J, Bo Y X, et al. Aeromonas hydrophila induces intestinal inflammation in grass carp (Ctenopharyngodon idella): An experimental model[J]. Aquaculture, 2014, 434: 171-178. doi: 10.1016/j.aquaculture.2014.08.015
    [32]
    Kotake S, Udagawa N, Takahashi N, et al. IL-17 in synovial fluids from patients with rheumatoid arthritis is a potent stimulator of osteoclastogenesis[J]. The Journal of Clinical Investigation, 1999, 103(9): 1345-1352. doi: 10.1172/JCI5703
    [33]
    Ding Y, Ao J Q, Chen X H. Comparative study of interleukin-17C (IL-17C) and IL-17D in large yellow croaker Larimichthys crocea reveals their similar but differential functional activity[J]. Developmental & Comparative Immunology, 2017, 76: 34-44.
    [34]
    张磊, 沈泽恩, 李红霞, 等. 鲤IL-17N的基因克隆、表达及促炎作用[J]. 中国水产科学, 2020, 27(12): 1402-1414.

    Zhang L, Shen Z E, Li H X, et al. Cloning, expression and pro-inflammatory effect of interleukin-17N in Cyprinus carpio[J]. Journal of Fishery Sciences of China, 2020, 27(12): 1402-1414 (in Chinese).
    [35]
    Xu P, Zhang X F, Wang X M, et al. Genome sequence and genetic diversity of the common carp, Cyprinus carpio[J]. Nature Genetics, 2014, 46(11): 1212-1219. doi: 10.1038/ng.3098
    [36]
    Louis A, Muffato M, Crollius H R. Genomicus: five genome browsers for comparative genomics in eukaryota[J]. Nucleic Acids Research, 2013, 41(D1): D700-D705.
    [37]
    Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2−△△C T method[J]. Methods, 2001, 25(4): 402-408. doi: 10.1006/meth.2001.1262
    [38]
    沈泽恩. 鲤鱼IL-17家族成员及其受体基因序列分析和表达特征[D]. 南京: 南京农业大学, 2019: 13-23.

    Shen Z E. Sequence analysis and expression characteristics of IL-17 family members and their receptor genes in Cyprinus carpio[D]. Nanjing: Nanjing Agricultural University, 2019: 13-23 (in Chinese).
    [39]
    Kim H S, Hwang D S, Jeong C B, et al. Identification and conservation of gene loss events of Hox gene clusters in the marine medaka (Oryzias melastigma)[J]. Journal of Experimental Zoology Part B:Molecular and Developmental Evolution, 2016, 326(7): 387-393. doi: 10.1002/jez.b.22713
    [40]
    You Z B, DuRaine G, Tien J Y L, et al. Expression of interleukin-17B in mouse embryonic limb buds and regulation by BMP-7 and bFGF[J]. Biochemical and Biophysical Research Communications, 2005, 326(3): 624-631. doi: 10.1016/j.bbrc.2004.11.087
    [41]
    Bie Q L, Jin C Q, Zhang B, et al. IL-17B: a new area of study in the IL-17 family[J]. Molecular Immunology, 2017, 90: 50-56. doi: 10.1016/j.molimm.2017.07.004
    [42]
    Lee J, Ho W H, Maruoka M, et al. IL-17E, a novel proinflammatory ligand for the IL-17 receptor homolog IL-17Rh1[J]. Journal of Biological Chemistry, 2001, 276(2): 1660-1664. doi: 10.1074/jbc.M008289200
    [43]
    Hoang C T, Hong Y, Truong A D, et al. Molecular cloning of chicken interleukin-17B, which induces proinflammatory cytokines through activation of the NF-κB signaling pathway[J]. Developmental and Comparative Immunology, 2017, 74: 40-48. doi: 10.1016/j.dci.2017.04.010
    [44]
    肖兰莹. 草鱼IL-17家族基因的克隆及其在肠道炎症中的功能鉴定[D]. 苏州: 苏州大学, 2018: 60-68.

    Xiao L Y. Grass Carp IL-17 Family genes: cloning, characterization and expression analysis during Aeromonas hydrophila-induced intestinal inflammation[D]. Suzhou: Suzhou University, 2018: 60-68 (in Chinese).
    [45]
    Hoesel B, Schmid J A. The complexity of NF-κB signaling in inflammation and cancer[J]. Molecular Cancer, 2013, 12(1): 86. doi: 10.1186/1476-4598-12-86
    • Related Articles

Catalog

    Get Citation
    PDF in Chinese
    XML
    Article views (344) PDF downloads (14) Cited by()
    Related

    Website Copyright © Editorial Office of Journal of Fisheries of China

    Address：No. 999, Hucheng Ring Road, Pudong New Area, Shanghai

    Tel：021-61900228 E-mail：jfc@china-fishery.com   

    京ICP备12043220号

    Supported by: Beijing Renhe Information Technology Co. Ltd  support: info@rhhz.net

    /

    DownLoad:  Full-Size Img  PowerPoint
    Return
    Return