我在看这篇 Herpes simplex virus disrupts NF-κB regulation by blocking its recruitment on the IκBα promoter and directing the factor on viral genes. 出处: THE JOURNAL OF BIOLOGICAL CHEMISTRY 2006, 281(11): 7110–7117 作者: Carla Amici, Antonio Rossi, Antonio Costanzo, Stefania Ciafre, Barbara Marinari, Mirna Balsamo, Massimo Levrero, and M. Gabriella Santoro (不好意思啊, 为了打字方便, 直接把kappa--> k; alpha--> a) 这篇论文主要探讨病毒感染後, 细胞NF-kB上升过程中可能的原因. 论文前几个实验已经推论几个时间点: 病毒感染--> IKK活性上升(感染後零小时及第三个小时)--> IkBa降解, 且NF-kB在感染後期不会被recruit到IkBa promoter(使IkBa产生, 回馈抑制NF-kB活性), 而会被recruit到病毒基因(ICP0)的promoter. 问题来了, 实验中将ICP0 promoter放到实验细胞株(HaCaT cell)中, 想要知道NF-kB被recruit到viral或cellular DNA, 可是我看了好久都看不懂. 原本我以为实验是要看病毒基因是否有嵌入宿主细胞的基因里, 但是老师说, 似乎这种病毒(HSV-1)不会嵌入宿主细胞.. 我不晓得这个实验的目的为何, 麻烦请病毒达人指点指点一下, 谢谢 :) 在此附上实验的原文: Viral ICP0 Promoter Shows Remarkable Avidity for NF-kB To investigate whether the differences observed in NF-kB recruitment could be a consequence of the different status of viral and cellular DNA organization (episomal versus chromosomal), we have generated HaCaT cells (HaCaT-ICP0-Luc cells) in which the ICP0 promoter controlling the expression of the luciferase reporter gene is stably integrated into the chromatin structure. HaCaT-ICP0-Luc cells were infected with HSV-1, and, at different times post-infection, were processed for luciferase or ChIP analysis. As shown in Fig. 5B, HSV-1 infection induces luciferase activity in these cells, indicating that the ICP0-Luc promoter is transcriptionally activated by the virus. In the same experiment, NF-kB recruitment to the integrated and to the free viral ICP0 promoters was analyzed at the end of the virus adsorption period and at 5 h post-infection, which correspond to the first and second waves of NF-kB activation, respectively. To discriminate the integrated form of the ICP0 promoter from the non-integrated viral promoter in the ChIP analysis, we have utilized the same upstream primer but different downstream primers (Fig. 5A). As shown in Fig. 5C, at the end of the 1-h adsorption period, NF-B is recruited to the IB promoter and to both forms (viral and integrated form) of the ICP0 promoter, indicating that during the first wave of NF-kB activation all the promoters analyzed behave similarly in respect to NF-kB recruitment. As expected, at 5 h p.i. NF-kB was not recruited to the IB promoter. Interestingly, at this time, NF-kB was recruited selectively to the viral form of the ICP0 promoter. When integrated into the host chromatin structure, the ICP0 promoter looses its ability to recruit the nuclear factor, behaving like IB (Fig. 5C). These results suggest that the differences in NF-kB recruitment observed between ICP0 and IB promoters could be due to differences in the status of DNA and/or to general chromatin modifications induced by HSV-1 during infection. --

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