Transdominant mutants of I kappa B alpha block Tat tumor necrosis factor synergistic activation of human immunodeficiency virus type 1 gene expression and virus multiplication
Transdominant mutants of I kappa B alpha block Tat tumor necrosis factor synergistic activation of human immunodeficiency virus type 1 gene expression and virus multiplication
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DOI：
10.1128/jvi.70.9.5777-5785.1996
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发表时间：
1996-09-01
影响因子：
5.4
通讯作者：
Hiscott, J
中科院分区：
文献类型：
--
作者：
Beauparlant, P;Kwon, H;Hiscott, J
人类免疫缺陷病毒1型（HIV-1）长末端重复序列（LTR）含有转录因子NF-κ B/Rel家族的两个结合位点，所述转录因子是通过炎性细胞因子如肿瘤坏死因子α（TNF-α）和白细胞介素-1转录激活病毒基因所需的。在本研究中，我们检测了I κ B α的反式显性突变体对TNF-α和HIV-1反式激活因子达特在Jurkat T细胞中协同激活HIV-1 LTR的作用。HIV-1 LTR驱动的基因表达的协同诱导代表了50至70倍的刺激，并且需要完整的HIV-1增强子和Tat-TAR元件相互作用，因为达特蛋白的突变（R52 Q，R53 Q）或TAR元件的凸出区域中消除达特与TAR结合的I κ B α的共表达不能刺激LTR表达，I κ B α的共表达以剂量依赖性方式抑制Tat-TNF-α对HIV LTR的激活，I κ B α的反式显性形式，在诱导剂介导的细胞凋亡所需的关键丝氨酸或苏氨酸残基中突变（S32 A，S36 A）和/或组成型检测I κ B α的磷酸化（S283 A、T291 A、T299 A）阻断HIV-1 LTR反式激活的能力，在诱导剂介导的刺激（t（1/2），>4 h）后，在N-末端位点突变的I κ B α分子不降解，并且能够有效地阻断HIV-1 LTR反式激活。引人注目的是，I κ B α（S32 A，S36 A）反式显性突变体在抑制HIV-1 LTR的协同诱导方面至少是野生型I κ B α的5倍。该突变体还有效地抑制HIV-1增殖在Cos-1细胞的单周期感染模型中，如通过病毒mRNA种类和病毒蛋白质产生的北方（RNA）印迹分析所测量的。这些实验提示了一种可能通过干扰NF-κ B/Rel信号通路来抑制HIV-1基因表达的策略。
The human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) contains two binding sites for the NF-kappa B/Rel family of transcription factors which are required for the transcriptional activation of viral genes by inflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) and interleukin-1. In the present study, we examined the effect of transdominant mutants of I kappa B alpha on the synergistic activation of the HIV-1 LTR by TNF-alpha and the HIV-1 transactivator, Tat, in Jurkat T cells, The synergistic induction of HIV-1 LTR-driven gene expression represented a 50- to 70-fold stimulation and required both an intact HIV-1 enhancer and Tat-TAR element interaction, since mutations in Tat protein (R52Q, R53Q) or in the bulge region of the TAR element that eliminated Tat binding to TAR were unable to stimulate LTR expression, Coexpression of I kappa B alpha inhibited Tat-TNF-alpha activation of HIV LTR in a dose-dependent manner, Transdominant forms of I kappa B alpha, mutated in critical serine or threonine residues required for inducer-mediated (S32A, S36A) and/or constitutive (S283A, T291A, T299A) phosphorylation of I kappa B alpha were tested for their capacity to block HIV-1 LTR transactivation, I kappa B alpha molecules mutated in the N-terminal sites were not degraded following inducer-mediated stimulation (t(1/2), >4 h) and were able to efficiently block HIV-1 LTR transactivation. Strikingly, the I kappa B alpha (S32A, S36A) transdominant mutant was at least five times as effective as wild-type I kappa B alpha in inhibiting synergistic induction of the HIV-1 LTR. This mutant also effectively inhibited HIV-1 multiplication in a single-cycle infection model in Cos-1 cells, as measured by Northern (RNA) blot analysis of viral mRNA species and viral protein production. These experiments suggest a strategy that may contribute to inhibition of HIV-1 gene expression by interfering with the NF-kappa B/Rel signaling pathway.