TBK1 of black carp plays an important role in host innate immune response against SVCV and GCRV.
TBK1 of black carp plays an important role in host innate immune response against SVCV and GCRV.
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DOI：
10.1016/j.fsi.2017.08.016
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发表时间：
2017-10
影响因子：
4.7
通讯作者：
Chuanzhe Yan;Jun Xiao;Jun Li;Hui Chen;J. Liu;Chanyuan Wang;C. Feng;Hao Feng
中科院分区：
文献类型：
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作者：
Chuanzhe Yan;Jun Xiao;Jun Li;Hui Chen;J. Liu;Chanyuan Wang;C. Feng;Hao Feng
TANK-BINDING KEK1(TANK-BINDING KEK1)在高等脊椎动物I型IFN的诱导中起关键作用。为了探讨TBK1在硬骨鱼中的功能，本研究克隆了青鱼(Mylophyngodon Piceus)的TBK1，并对其进行了鉴定。青鱼TBK1基因全长由2857个核苷酸组成，编码的bcTBK1蛋白由727个氨基酸组成，包括一个N-末端的激酶结构域(KD)、一个泛素样结构域(ULD)和两个C-末端的卷曲。在所有组织中均检测到bcTBK1的转录，除GCRV侵袭后的肌肉组织外，所有组织中的bcTBK1基因表达水平在SVCV或GCRV感染后均升高。SVCV、GCRV或Poly(I：C)可上调Mylophyngodon Piceusfin(MPF)细胞中bcTBK1的转录，但不受内毒素刺激。免疫印迹法检测bcTBK1迁移约80 kDa，免疫荧光染色鉴定为胞浆蛋白。在报告实验中，bcTBK1具有较强的干扰素诱导能力，并在EPC细胞中对GCRV和SVCV具有较强的抗病毒活性。报告实验表明，青鱼TRAF6(BcTRAF6)上调了bcTBK1诱导的干扰素的表达，并且bcTBK1和bcTRAF6蛋白在EPC细胞中共表达时，两者的亚细胞分布重叠。综上所述，本研究支持bcTBK1在青鱼抗SVCV和GCRV天然免疫应答中的重要作用，其活性受bcTRAF6正向调节。
Tank-binding kinase 1 (TBK1) plays a pivotal role in the induction of type I IFNs in higher vertebrates. To explore the function of TBK1 in teleost, TBK1 of black carp (Mylopharyngodon Piceus) was cloned and characterized in this paper. The full-length cDNA of black carp TBK1 (bcTBK1) consists of 2857 nucleotides and the predicted bcTBK1 protein contains 727 amino acids, which includes an N-terminal kinase domain (KD), an ubiquitin-like domain (ULD) and two C-terminal coiled-coils. The transcription of bcTBK1 was constitutively detected in all the selected tissues and bcTBK1 mRNA level was increased in all selected tissues in response to SVCV or GCRV infection except that in muscle post GCRV invasion. The transcription of bcTBK1 inMylopharyngodon Piceusfin (MPF) cells was up-regulated by the stimulation of SVCV, GCRV or poly (I:C) but not by LPS treatment. bcTBK1 migrated around 80 kDa in immunoblot assay and was identified as a cytosolic protein by immunofluorescence staining. bcTBK1 showed strong IFN-inducing ability in reporter assay and presented strong antiviral activity against both GCRV and SVCV in EPC cells. The reporter assay demonstrated that TRAF6 of black carp (bcTRAF6) up-regulated bcTBK1-induced IFN expression and the subcellular distribution of bcTBK1 overlapped with that of bcTRAF6 when these two proteins were co-expressed in EPC cells. Taken together, our study support the conclusion that bcTBK1 plays an important role in the antiviral innate immune response of black carp against SVCV and GCRV, in which its activity was positively regulated by bcTRAF6.