Construction of HEK-293T Cell Line with Stable Expression of Histone H2B and Evaluation of Proliferation Efficiency of Herpes simplex virus
WEI Da-Li1,2, QU Min2, KE Qun-Hua1,2, LIU Yi-Jia2, ZHANG Li-Na1,2, SUN Yue-Feng1,2, YIN Xiang-Ping2, JI Peng1,*
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China; 2 Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences/College of Veterinary Medicine, Lanzhou University/State Key Laboratory for Animal Disease Controland Prevention/College of Veterinary Medicine, Lanzhou 730046, China
Abstract：Histone H2B is an important part of the nucleosome, which affects gene transcription and biological processes, and is expected to become a molecular target for disease treatment. In this study, lentiviral transfection was applied to construct an HEK-293T cell line with stable expression of histone H2B. The recombinant plasmid pLVX-mCherry-puro-H2B was constructed using the H2B gene as the target and then co-transfected with auxiliary plasmids pMD2.G and psPAX2 into HEK-293T cells for lentiviral packaging. The Lentivirus was transduced to HEK-293T cells, and a stable histone H2B-expressing cell line was obtained through purinomycin pressure screening, which was named HEK-293T-H2B cell line. The replication capacity of Herpes simplex virus (HSV) in the HEK-293T-H2B cell line was determined by qPCR and Western blot assays. The results showed that the replication of HSV was significantly weakened in the HEK-293T-H2B cell line. qPCR was used to detect the expression of downstream interferon genes in HEK-293T-H2B and wild-type HEK-293T cells infected with HSV. The results showed that the HEK-293T-H2B cell line promoted the expression of genes induced by HSV downstream typeⅠinterferon. In conclusion, the HEK-293T-H2B cell line constructed in this study can significantly prompte the typeⅠinterferon signaling pathway and inhibit the proliferation of HSV, which provides a basis for further research on the antiviral function of histone H2B.
魏达礼, 瞿敏, 柯群华, 刘艺佳, 张丽娜, 孙跃峰, 殷相平, 纪鹏. 稳定表达组蛋白H2B的HEK-293T细胞系构建及单纯疱疹病毒增殖效率评价[J]. 农业生物技术学报, 2026, 34(1): 221-228.
WEI Da-Li, QU Min, KE Qun-Hua, LIU Yi-Jia, ZHANG Li-Na, SUN Yue-Feng, YIN Xiang-Ping, JI Peng. Construction of HEK-293T Cell Line with Stable Expression of Histone H2B and Evaluation of Proliferation Efficiency of Herpes simplex virus. 农业生物技术学报, 2026, 34(1): 221-228.
[1] 刘伟, 李梦娇, 郭佩东, 等. 2021.稳定表达人TMPRSS2基因BHK细胞系的建立及其对新城疫弱毒增殖效率评价[J].中国兽医科学, 51(05):594-600. (Liu W, LI M J, Guo P D, et al.2021. Establishment of BHK cell line stably expressing human TMPRSS2 gene and evaluation of lentogenic Newcastle disease virus proliferation efficiency[J]. China Veterinary Science, 51(05): 594-600.) [2] 王佳豪,冯利杰,张瑶,等.2024.稳定表达PSMB9-eGFP-His蛋白的THP-1细胞系建立及免疫蛋白酶体活性检测[J].生物工程学报, 40(07): 2282-2293. (Wang J H, Feng L J, Zhang Y, et al.2024. Establishment of a stable THP-1 cell line expressing PSMB9-eGFP-His protein and detection of immunoproteasome activity[J]. Chinese Journal of Biotechnology, 40(7): 2282-2293.) [3] Arena T A, Harms P D, Wong A W.2018. High throughput transfection of HEK293 cells for transient protein production[J]. Methods in Molecular Biology, 1850: 179-187. [4] Bai L, Xu J, Zeng L, et al.2024. A review of HSV pathogenesis, vaccine development, and advanced applications[J]. Molecular Biomedicine, 5(1): 35. [5] Bannister A J, Kouzarides T.2011. Regulation of chromatin by histone modifications[J]. Cell Research, 21(3): 381-395. [6] Chen J, Lu Z, Gong W, et al.2022. Epstein-Barr virus protein BKRF4 restricts nucleosome assembly to suppress host antiviral responses[J]. Proceedings of the National Academy of Sciences of the USA, 119(37): e2203782119. [7] Conceição-Silva F, Reis C S M, De Luca P M, et al.2021. The immune system throws its traps: Cells and their extracellular traps in disease and protection[J]. Cells, 10(8): 1891. [8] Fu H, Pan D.2024. Mechanisms of HSV gene regulation during latency and reactivation[J]. Virology, 110324. [9] Honda K, Taniguchi T.2006. IRFs: Master regulators of signalling by Toll-like receptors and cytosolic pattern-recognition receptors[J]. Nature Reviews Immunology, 6(9): 644-658. [10] Jiang L, Smith J N, Anderson S L, et al.2007. Global assessment of combinatorial post-translational modification of core histones in yeast using contemporary mass spectrometry: LYS4 trimethylation correlates with degree of acetylation on the same H3 tail[J]. Journal of Biological Chemistry, 282(38): 27923-27934. [11] Khadka J, Pesok A, Grafi G.2020. Plant histone HTB (H2B) variants in regulating chromatin structure and function[J]. Plants, 9(11): 1435. [12] Li J, Zhang J, Sun P, et al.2024. Porcine reproductive and respiratory syndrome virus nonstructural protein 2 promotes the autophagic degradation of adaptor protein SH3KBP1 to antagonize host innate immune responses by enhancing K63-linked polyubiquitination of RIG-I[J]. PLOS Pathogens, 20(10): e1012670. [13] Li X, Ye Y, Peng K, et al.2022. Histones: The critical players in innate immunity[J]. Frontiers in Immunology, 13: 1030610. [14] Lin Y, Qiu T, Wei G, et al.2022. Role of histone post-translational modifications in inflammatory diseases[J]. Frontiers in Immunology, 13: 852272. [15] Liu H, Li C, He W, et al.2022. Free ISG15 inhibits Pseudorabies virus infection by positively regulating typeⅠIFN signaling[J]. PLOS Pathogens, 18(10): e1010921. [16] Parra M A, Kerr D, Fahy D, et al.2006. Deciphering the roles of the histone H2B N-terminal domain in genome-wide transcription[J]. Molecular and Cellular Biology, 26(10): 3842-3852. [17] Quina A S, Buschbeck M, Di Croce L.2006. Chromatin structure and epigenetics[J]. Biochemical Pharmacology, 72(11): 1563-1569. [18] Wyrick J J, Parra M A.2009. The role of histone H2A and H2B post-translational modifications in transcription: A genomic perspective[J]. Biochimica et Biophysica Acta (BBA)-Gene Regulatory Mechanisms, 1789(1): 37-44. [19] Savidis G, Perreira J M, Portmann J M, et al.2016. The IFITMs inhibit Zika virus replication[J]. Cell Reports, 15(11): 2323-2330. [20] Schoggins J W, Wilson S J, Panis M, et al.2011. A diverse range of gene products are effectors of the typeⅠinterferon antiviral response[J]. Nature, 472(7344): 481-485. [21] Xu H, Wu M, Ma X, et al.2021. Function and mechanism of novel histone posttranslational modifications in health and disease[J]. BioMed Research International, 2021(1): 6635225. [22] Zhu Z, Li D, Jia Z, et al.2023. Global histone H2B degradation regulates insulin/IGF signaling‐mediated nutrient stress[J]. The EMBO Journal, 42(19): e113328.
