Acquisition of HIV-1 Resistance in T Lymphocytes Using an ACA-Specific E. coli mRNA Interferase
Acquisition of HIV-1 Resistance in T Lymphocytes Using an ACA-Specific E. coli mRNA Interferase
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DOI：
10.1089/hum.2010.001
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发表时间：
2011-01-01
影响因子：
4.2
通讯作者：
Kato, Ikunoshin
中科院分区：
文献类型：
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作者：
Chono, Hideto;Matsumoto, Kazuya;Kato, Ikunoshin
HIV-1长末端重复序列(LTR)控制的基因转录激活需要反式激活反应元件(TAR)和TAT蛋白。缺乏功能性Tat基因的HIV-1突变株不能增殖。在这里，我们采取了一种基因方法来抑制HIV-1的复制，该方法基于TAT依赖的MazF的产生，MazF是一种来自大肠杆菌的ACA特异性内切核酸酶(mRNA干扰酶)。当被诱导时，MazF被认为能引起哺乳动物细胞中Bak和NBK依赖性的细胞死亡。我们首先构建了一个逆转录病毒载体，将mazF(ACA-less)基因插入到HIV-1 LTR的控制下，然后将其导入CD4+T淋巴样CEM-SS细胞，在HIV-1感染时，诱导mazF基因破坏感染的HIV-1mRNA，防止HIV-1复制。事实上，当转导细胞感染HIV-1 IIIB时，病毒复制被有效地抑制，因为在培养基中检测不到HIV-1 IIIB p24。始终如一的是，不仅细胞生长，而且CD4水平都没有受到感染的影响。这些结果表明，HIV-1-LTR调控的mazF基因在HIV-1 IIIB感染后被有效诱导，足以破坏感染HIV-1 IIIB的病毒mRNA，完全阻止病毒在细胞中的增殖，但不影响正常细胞的生长。这些结果表明，转导了HIV-1-LTRMAZF基因的T细胞获得了对HIV-1的抵抗力，这为将HIV-1-LTRMAZF基因用于抗HIV基因治疗提供了诱人的潜力。
Transcriptional activation of gene expression directed by the long terminal repeat (LTR) of HIV-1 requires both the transactivation response element (TAR) and Tat protein. HIV-1 mutants lacking a functional tat gene are not able to proliferate. Here we take a genetic approach to suppress HIV-1 replication based on Tat-dependent production of MazF, an ACA-specific endoribonuclease (mRNA interferase) from Escherichia coli. When induced, MazF is known to cause Bak- and NBK-dependent apoptotic cell death in mammalian cells. We first constructed a retroviral vector, in which the mazF (ACA-less) gene was inserted under the control of the HIV-1 LTR, which was then transduced into CD4+ T-lymphoid CEM-SS cells in such a way that, upon HIV-1 infection, the mazF gene is induced to destroy the infecting HIV-1 mRNA, preventing HIV-1 replication. Indeed, when the transduced cells were infected with HIV-1 IIIB, the viral replication was effectively inhibited, as HIV-1 IIIB p24 could not be detected in the culture medium. Consistently, not only cell growth but also the CD4 level was not affected by the infection. These results suggest that the HIV-1-LTR-regulated mazF gene was effectively induced upon HIV-1 IIIB infection, which is sufficient enough to destroy the viral mRNA from the infected HIV-1 IIIB to completely block viral proliferation in the cells, but not to affect normal cell growth. These results indicate that the T cells transduced with the HIV-1-LTR-regulated mazF gene acquire HIV-1 resistance, providing an intriguing potential for the use of the HIV-1-LTR-regulated mazF gene in anti-HIV gene therapy.