Regulation Mechanisms of HIV LTR Demethylation through Reactivation Signaling

HIV LTR 去甲基化通过再激活信号的调节机制

• 批准号: 12670274
• 负责人: ISHIDA Takaomi
• 金额: $ 0.9万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670274/
• 关键词: HIV; LTR; Transcription; CpG Methylation; Latent Infection; LTR; 転写抑制; 潜伏感染; Histone Code; クロマチン免疫沈降法; ヌクレオソーム; 潜伏HIV再活性化シグナルによるLTRの脱メチル化制御; 細胞内シグナル伝達

Introduction of potent anti-HIV combination chemotherapy has focused reseacher's interest on the mechanisms of viral latency, because eradication of HIV cannot be achieved without eradication of latent HIV in reservoir pool. CpG methylation has long been impricated in the viral latency as one base for repression of gene expression. We could find that the levels of CpG methylation were inversly correlated with the viral gene expression in chironically infected cell lines. Induction of viral gene expresson by TNF-α in ACH-2 cell line was shown to be associated with demethylation of CpG sequence in the provirus LTR.Because we could find that 5'-LTR specific methylation was occurred in the HTLV-1 provirus LTR, we designed the bisulfite modified PCR sense primer located in the 5'-flanking sequence of the integrated provirus and that in the tat coding region and selectively methylation status analyzed. Results revealed selective hypermethylation of 5'-LTR and hypomethylation of 3'-LTR in the ACH-2 cell line. The same selectivity was observed in the chilonically infected cell lines, ATL cases and HTLV-1 asymptomatic carriers. It was possible that mammalian cells has the ability to hypermethylate the integrated retrovirus 5'-LTR or to demethylate the 3'-LTR.

有效的抗 HIV 联合化疗的引入使研究人员的兴趣集中在病毒潜伏期的机制上，因为如果不消除病毒潜伏期中的潜伏期病毒，就无法消灭 HIV，而 CpG 甲基化长期以来一直被认为是病毒潜伏期的一种抑制基础。我们可以发现，CpG 甲基化水平与 TNF-α 诱导的病毒基因表达呈负相关。 ACH-2细胞系被证明与原病毒LTR中CpG序列的去甲基化有关。由于我们发现HTLV-1原病毒LTR中发生了5'-LTR特异性甲基化，因此我们设计了亚硫酸氢盐修饰的PCR正义引物，位于整合原病毒的 5'-侧翼序列和 tat 编码区的选择性甲基化状态分析结果显示 5'-LTR 的选择性高甲基化和低甲基化。在 ACH-2 细胞系中观察到 3'-LTR 的选择性在慢性感染细胞系、ATL 病例和 HTLV-1 无症状携带者中观察到，哺乳动物细胞可能具有使整合的逆转录病毒 5' 高度甲基化的能力。 -LTR 或使 3'-LTR 去甲基化。

项目成果

Tanaka.T., et al.: "Latent HIV-1 reactivation in transgenic mice requires cell cycle-dependent demethylation of CREB/ATF sites in the LTR"AIDS. 17. 165-175 (2002)

Tanaka.T.等人：“转基因小鼠中潜在的HIV-1重新激活需要LTR中CREB/ATF位点的细胞周期依赖性去甲基化”AIDS。

Koiwa.T., et al.: "5'-Long Terminal Repeat-Selective CpG Methylation of Latent Human T-Cell Leukemia Virus Type 1 Provirus In Vitro and In Vivo"Journal of Virology. 76. 9389-9397 (2002)

Koiwa.T.等人：“体外和体内潜伏人类T细胞白血病病毒1型原病毒的5-长末端重复选择性CpG甲基化”病毒学杂志。

Horie, R., et al.: "Ligand-independent signaling by overexpressed CD3O drives NF-kB activation in Hodgkin-Reed starnberg cells"Oncogene. (in press). (2002)

Horie, R., 等人：“过表达的 CD3O 驱动 NF-kB 激活的配体独立信号传导霍奇金-里德斯塔恩伯格细胞”癌基因。

Tanaka J., Ishida T., Choi B.I., Yasuda J., Watanabe T., Iwakura Y.: "Latent HIV-1 reactivation in transgenic mice requires cell cycle-dependent demethylation of CREB/ATF sites in the LTR"AIDS.. 17. 165-175 (2002)

Tanaka J.、Ishida T.、Choi B.I.、Yasuda J.、Watanabe T.、Iwakura Y.：“转基因小鼠中潜在的 HIV-1 重新激活需要 LTR 中 CREB/ATF 位点的细胞周期依赖性去甲基化”AIDS..

Koiwa T., Hamano-Usami A., Ishida T., Okayama A., Yamaguchi K., Watanabe T.: "5'-LTR-selective CpG methylation of latency infected HTLV-1 provirus in vitro and in vivo"Journal of Virology. 76. 9389-9397 (2002)

Koiwa T.、Hamano-Usami A.、Ishida T.、Okayama A.、Yamaguchi K.、Watanabe T.：“体外和体内潜伏感染的 HTLV-1 原病毒的 5-LTR 选择性 CpG 甲基化”杂志