| 170 | 0 | 171 |
| 下载次数 | 被引频次 | 阅读次数 |
本研究以痘苗病毒天坛株（Vaccinia Virus Tian Tan Strain,VTT）为对象，旨在明确N1L基因的表达调控序列。我们克隆了N1L起始密码子ATG上游不同长度的调控序列片段，构建了表达载体，并利用报告基因萤火虫荧光素酶（Firefly luciferase,Fluc）的表达来确定N1L表达调控序列区域。通过PROMO在线预测网站，预测了与调控序列结合的转录因子。结果显示，VTT中N1L基因的表达受到上游调控序列的调控，其中以-404bp起始的片段启动的Fluc表达量最高。PROMO预测表明该区域存在多个转录因子结合位点。本研究为痘苗病毒基因表达特点的研究提供了实验依据，并为N1L基因功能及机制研究奠定了基础。
Abstract：We aimed to elucidate the regulatory sequences governing expression of N1L in vaccinia virus Tian Tan strain. Various lengths of regulatory sequence fragments upstream of the N1L start codon ATG were cloned, and expression vectors were constructed. Expression of the reporter gene Firefly luciferase(Fluc) served to identify regions of high N1L expression in regulatory sequences. Utilizing an online prediction tool(PROMO), predictions were made regarding transcription factors binding to regulatory sequences. N1L expression in VTT was regulated by upstream regulatory sequences, with the fragment commencing at-404bp initiating the highest Fluc expression. PROMO predictions indicated the presence of multiple binding sites in transcription factors within this region. This study furnishes experimental evidence for understanding genetic expression in the vaccinia virus, laying the groundwork for further research into the function and mechanism of N1L.
[1] Li M, Zhang M, Ye Q, et al. Preclinical and clinical trials of oncolytic vaccinia virus in cancer immunotherapy:a comprehensive review[J]. Cancer Biol Med, 2023, 20(9):646-661. DOI:10. 20892/j.issn. 2095-3941. 2023. 0202.
[2] Wang Y, Griffiths A, Brackney D E, et al. Generation of multiple Arbovirus-like particles using a rapid recombinant vaccinia virus expression platform[J].Pathogens, 2022, 11(12):1505. DOI:10. 3390/pathogens11121505.
[3] Shen X, Basu R, Sawant S, et al. HIV-1 gp120 and modified vaccinia virus ankara(MVA)gp140 boost immunogens increase immunogenicity of a DNA/MVA HIV-1 Vaccine[J]. J Virol, 2017, 91(24):e01077-17.DOI:10. 1128/jvi. 01077-17.
[4] Wei C M, Moss B. Methylated nucleotides block 5'-terminus of vaccinia virus messenger RNA[J]. Proc Natl Acad Sci U S A, 1975, 72(1):318-322. DOI:10. 1073/pnas. 72. 1. 318.
[5] Kates J, Beeson J. Ribonucleic acid synthesis in vaccinia virus. I. The mechanism of synthesis and release of RNA in vaccinia cores[J]. J Mol Biol, 1970, 50(1):1-18. DOI:10. 1016/0022-2836(70)90100-2.
[6] Tombácz D, Prazsák I, Torma G, et al. Time-course transcriptome profiling of a poxvirus using long-read fulllength assay[J]. Pathogens, 2021, 10(8):919. DOI:10. 3390/pathogens10080919.
[7] Greseth M D, Traktman P. The life cycle of the vaccinia virus genome[J]. Annu Rev Virol, 2022, 9(1):239-259. DOI:10. 1146/annurev-virology-091919-104752.
[8] Yang Z, Bruno D P, Martens C A, et al. Genome-wide analysis of the 5'and 3'ends of vaccinia virus early mRNAs delineates regulatory sequences of annotated and anomalous transcripts[J]. J Virol, 2011, 85(12):5897-5909. DOI:10. 1128/jvi. 00428-11.
[9] Yang Z, Cao S, Martens C A, et al. Deciphering poxvirus gene expression by RNA sequencing and ribosome profiling[J]. J Virol, 2015, 89(13):6874-6886. DOI:10. 1128/jvi. 00528-15.
[10]阮力.痘苗病毒天坛株载体研究与应用概述[J].微生物与感染，2013, 8(1):2-8.
[11]Wollenberg A, Engler R. Smallpox, vaccination and adverse reactions to smallpox vaccine[J]. Curr Opin Allergy Clin Immunol, 2004, 4(4):271-275. DOI:10. 1097/01. all. 0000136758. 66442. 28.
[12]黄盼盼，复制缺陷型痘苗病毒NTV细胞生物学特性及复制缺陷机制初步探究[D]. 2020，中国疾病预防控制中心.
[13]Aoyagi M, Zhai D, Jin C, et al. Vaccinia virus N1L protein resembles a B cell lymphoma-2(Bcl-2)family protein[J]. Protein Sci, 2007, 16(1):118-124. DOI:10. 1110/ps. 062454707.
[14]Cooray S, Bahar M W, Abrescia N G A, et al.Functional and structural studies of the vaccinia virus virulence factor N1 reveal a Bcl-2-like anti-apoptotic protein[J]. J Gen Virol, 2007, 88(Pt 6):1656-1666.DOI:10. 1099/vir. 0. 82772-0.
[15]DiPerna G, Stack J, Bowie A G, et al. Poxvirus protein N1L targets the I-kappaB kinase complex,inhibits signaling to NF-kappaB by the tumor necrosis factor superfamily of receptors, and inhibits NF-kappaB and IRF3 signaling by toll-like receptors[J]. J Biol Chem, 2004, 279(35):36570-36578. DOI:10. 1074/jbc. M400567200.
[16]阮力，郑浩强，徐水婵，等.呼吸道合胞病毒糖蛋白F和G在同一个重组痘苗病毒中的表达[J].病毒学报，1992(02):101-109.
[17]Bartlett N, Symons J A, Tscharke D C, et al. The vaccinia virus N1L protein is an intracellular homodimer that promotes virulence[J]. J Gen Virol, 2002, 83(Pt8):1965-1976. DOI:10. 1099/0022-1317-83-8-1965.
[18]王娟，转录因子ⅡA及其识别元件在RNA聚合酶II指导的基因转录中的作用和调节机制的研究[D]. 2019，武汉科技大学.
[19]Calkhoven C F, Ab G. Multiple steps in the regulation of transcription-factor level and activity[J]. Biochem J,1996, 317(Pt 2)(Pt 2):329-342. DOI:10. 1042/bj3170329.
[20]Mishal R, Luna-Arias J P. Role of the TATA-box binding protein(TBP)and associated family members in transcription regulation[J]. Gene, 2022, 833:146581.DOI:10. 1016/j. gene. 2022. 146581.
基本信息:
DOI：10.13242/j.cnki.bingduxuebao.004519
中图分类号:R373
引用信息:
[1]翟玉倩,韩一泽,吴长城等.痘苗病毒天坛株N1L基因表达调控序列的克隆与特性分析[J].病毒学报,2024,40(03):484-491.DOI:10.13242/j.cnki.bingduxuebao.004519.
基金信息: