The 156KELK159 tetrapeptide of HIV-1 integrase is critical for lentiviral gene integration
The 156KELK159 tetrapeptide of HIV-1 integrase is critical for lentiviral gene integration
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HIV-1整合酶的156KELK159四肽对于慢病毒基因整合至关重要
DOI：
10.1007/s11033-011-0744-z
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发表时间：
2011-05
期刊：
影响因子：
--
通讯作者：
陈金中
中科院分区：
文献类型：
--
作者：
陈金中
HIV-1整合酶（HIV-1 IN）是HIV-1慢病毒载体的关键元件，是载体基因插入宿主基因组中得以稳定维持的关键。HIV-1 IN除整合外还具有多种功能，如参与病毒粒子形态学、病毒DNA合成和病毒DNA核输入等。在我们的研究中，酵母双杂交试验鉴定了HIV-1 IN中对HIV-1 IN和Daxx相互作用至关重要的四肽156 KELK 159。为了研究HIV-1 IN的四肽156 KELK 159的功能，将野生型HIV-1 IN和缺失156 KELK 159的突变体用于包装含有EGFP报告基因的慢病毒。基于p24的滴度测定揭示了删除四肽不影响病毒包装。用病毒特异性引物进行真实的实时定量PCR检测。但156 KELK 159对慢病毒基因整合至关重要。删除四肽使表达报告基因的细胞百分比显着下降，并且不影响进入细胞或细胞核的DNA水平。采用真实的时间反转录PCR和流式细胞术检测慢病毒报告基因在感染维持细胞中的表达，结果显示156 KELK 159对慢病毒载体基因的表达没有影响。本研究结果有助于阐明慢病毒基因整合的调控机制。
HIV-1 integrase (HIV-1 IN), a key element of HIV-1-derived lentiviral vectors, is crucial for the stable maintenance of the vector gene by inserting them into host genome. HIV-1 IN has been found to have functions other than integration, such as involving in virion morphology, viral DNA synthesis and viral DNA nuclear import. In our study, the yeast two-hybrid assay identified a tetrapeptide 156KELK159 in HIV-1 IN that was crucial for HIV-1 IN and Daxx interaction. To investigate the functions of the tetrapeptide 156KELK159 of the HIV-1 IN, both the wild type HIV-1 IN and a mutant without 156KELK159 were used to package the EGFP reporter gene contained lentivirus. p24 based titer assay revealed that deleting the tetrapeptide did not affect virus packaging. The result was verified by quantitative real time PCR with viral specific primers. But the 156KELK159 was crucial for lentiviral gene integration. Deleting the tetrapeptide made the percentage of cells expressing the reporter gene significantly decreased and did not affect the level of DNA entered into the cells or nucleus. Real time reverse transcription PCR and FACS were used to detect the lentiviral report gene expression in infection maintaining cells and revealed 156KELK159 did not affect lentiviral vector gene expression. Our results may shed light on the regulatory mechanism of gene integration of lentivirus.
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影响因子：
--
作者：
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通讯作者：
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通讯作者：
GOFF, SP
影响因子：
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作者：
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通讯作者：
Masuda, T