HTLV-1 TAX1 AS AN EXTRACELLULAR CYTOKINE

人类T细胞嗜淋巴病毒I型(HTLV-I)与两种病毒有关 人类疾病、成人T细胞白血病(ATL)和热带痉挛 截瘫/HTLV-I相关性脊髓病(TSP/HAM)。HTLV-I出租车1号 从受感染的淋巴细胞中释放出来，并发挥细胞外功能 刺激基因表达和增殖的细胞因子， 未感染的淋巴细胞。人T细胞嗜淋巴病毒I型 (HTLV-I)Tax1诱导血管内皮细胞活化和核定位 细胞转录因子，核因子-kappaB。对细胞的处理 蛋白激酶C(PKC)抑制剂Calphostin C阻断了 HTLV-I转化的C81细胞中核因子-kappaB DNA结合活性的研究 Tax1刺激的小鼠前B细胞，提示PKC是一种 在核因子-kappaB诱导途径中的重要中间体。确实是 进一步证明了Tax1与PKC结合并激活PKC。 PKC与表达Tax1的抗Tax1血清免疫共沉淀 存在外源紫杉醇1的MT4提取物和Jurkat提取物 蛋白。此外，谷胱甘肽-S-转移酶-Tax1蛋白 与阿尔法、德尔塔和ETAPKC同工酶特异性结合 在兔网织红细胞裂解物中合成。将税项1加入 体外激酶反应导致Tax1和An的磷酸化 PKC的自磷酸化增加了18倍。转染法 野生型Tax1刺激的Jurkat细胞膜转位的研究 PKC的基本原理。相比之下，ax1突变体M22未能刺激 核因子-kappaB依赖转录，不能刺激膜 PKC易位。Tax1不会直接增加PKC IkappaBalpha的磷酸化。我们的结果与一个 Tax1与PKC相互作用并刺激膜的模型 PKC通路的转位和触发。中的后续步骤 PKC级联可能刺激IkappaBalpha的磷酸化。 核因子-kappaB调控多种病毒和细胞基因的表达 包括HIV LTR、MHC I类和IL-2Rα细胞因子基因。它 已证实Rb刺激核因子-kappaB p50的结合 同源二聚体。在体外凝胶位移结合中加入Rb蛋白 实验刺激p50结合超过10倍。有趣的是，通过 分析核因子-kappaB依赖的体外转录活性，我们 证明Rb抑制p50的转录活性。 胰凝乳酶分析表明Rb诱导构象变化 在核因子-kappaB-DNA复合体中，导致结合a 转录不活跃的复合体。最后，我们通过以下方式进行演示 Rb-p50复合体存在的免疫共沉淀分析 Jurkat细胞提取物。我们的结果表明，Rb可能扮演着一种 在调节核因子-kappaB转录活性中的重要作用。

Human T-cell lymphotropic virus type I (HTLV-I) is associated with two human diseases, adult T-cell leukemia (ATL), and tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM). HTLV-I Tax1 is released from infected lymphocytes and functions as an extracellular cytokine to stimulate gene expression in, and proliferation of, uninfected lymphocytes. Human T-cell lymphotropic virus type I (HTLV-I) Tax1 induces the activation and nuclear localization of the cellular transcription factor, NF-kappaB. Treatment of cells with calphostin C, a protein kinase C (PKC) inhibitor, blocked induction of NF-kappaB DNA binding activity in HTLV-I-transformed C81 cells and Tax1-stimulated murine pre-B cells, suggesting that PKC was an important intermediate in the NF-kappaB induction pathway. It was further demonstrated that Tax1 associates with, and activates, PKC. PKC was coimmunoprecipitated with anti-Tax1 sera from Tax1-expressing MT4 extracts and Jurkat extracts in the presence of exogenous Tax1 protein. In addition, Glutathione-S-transferase (GST)-Tax1 protein bound specifically to the alpha, delta and eta PKC isoenzymes synthesized in rabbit reticulocyte lysates. The addition of Tax1 to in vitro kinase reactions leads to the phosphorylation of Tax1 and an eighteen-fold increase in the autophosphorylation of PKC. Transfection of Jurkat cells with wild-type Tax1 stimulated membrane translocation of PKC. In contrast, Tax1 mutant M22, which fails to stimulate NF-kappaB-dependant transcription, failed to stimulate membrane translocation of PKC. Tax1 did not directly increase PKC phosphorylation of IkappaBalpha. Our results are consistent with a model in which Tax1 interacts with PKC and stimulates membrane translocation and triggering of the PKC pathway. Subsequent steps in the PKC cascade likely stimulate phosphorylation of IkappaBalpha. NF-kappaB regulates expression of several viral and cellular genes including the HIV LTR, MHC class I and IL-2Ralpha cytokine genes. It has been demonstrated that Rb stimulates binding of the NF-kappaB p50 homodimer. Addition of Rb protein to an in vitro gel shift binding assay stimulated p50 binding greater than ten-fold. Interestingly, by analyzing NF-kappaB-dependent transcription activity in vitro, we demonstrate that Rb suppresses transcriptional activity of p50. Chymotrypsin analysis suggests that Rb induces a conformational change in the NF-kappaB-DNA complex resulting in binding of a transcriptionally inactive complex. Finally, we demonstrate by coimmunoprecipitation analysis that the Rb-p50 complex is present in Jurkat cell extracts. Our results suggest that Rb may play an important role in regulation of NF-kappaB transcriptional activity.