Regulation of the Epstein-Barr virus C promoter by AUF1 and the cyclic AMP/protein kinase A signaling pathway
Regulation of the Epstein-Barr virus C promoter by AUF1 and the cyclic AMP/protein kinase A signaling pathway
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DOI：
10.1128/jvi.74.17.8166-8175.2000
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发表时间：
2000-09-01
影响因子：
5.4
通讯作者：
Ling, PD
中科院分区：
文献类型：
--
作者：
Fuentes-Panan치, EM;Peng, RS;Ling, PD
EBNA 2是EB病毒（EBV）编码的蛋白质，其调节EBV驱动的B细胞永生化所需的病毒和细胞基因的表达。阐明EBNA 2调节病毒和细胞基因表达的机制对于理解人类中EBV诱导的B细胞永生化和病毒潜伏期是必要的。EBNA 2通过与细胞DNA结合蛋白CBF 1（RBPJk）相互作用靶向潜伏C启动子（Cp）。Cp中的EBNA 2增强子还结合另一种细胞因子，C启动子结合因子2（CBF 2），其蛋白产物尚未鉴定。在Cp中的EBNA 2增强子内，我们先前已经鉴定了CBF 2结合所需的DNA序列，并且还确定了该元件是EBNA 2有效激活Cp所需的。在这项研究中，CBF 2活性的生化纯化和微测序。测序的肽与hnRNP蛋白AUF 1相同。在凝胶迁移率变动分析中，针对AUF 1的抗体而不是针对相关hnRNP蛋白的抗体与CBF 2反应。此外，细胞环AMP（cAMP）/蛋白激酶A（PKA）信号转导途径的刺激导致从刺激的细胞提取的可检测的CBF 2/AUF 1结合活性增加。此外，CBF 2结合位点能够赋予EBNA 2对异源启动子的响应性，当转染细胞用激活PKA的化合物处理或通过共转染表达PKA的组成型活性催化亚基的质粒时，EBNA 2介导的潜伏期Cp的刺激也在类似的共转染测定中增加。这些结果进一步支持了CBF 2在介导EBNA 2反式激活中的重要作用;它们将hnRNP蛋白AUF 1鉴定为CBF 2的主要组分，并且也是除了CBF 1结合元件之外的顺式作用序列的第一个证据，该顺式作用序列能够赋予对EBNA 2的响应性。
EBNA2 is an Epstein-Barr virus (EBV)-encoded protein that regulates the expression of viral and cellular genes required for EBV-driven B-cell immortalization. Elucidating the mechanisms by which EBNA2 regulates viral and cellular gene expression is necessary to understand EBV-induced B cell immortalization and viral latency in humans. EBNA2 targets to the latency C promoter (Cp) through an interaction with the cellular DNA binding protein CBF1 (RBPJk). The EBNA2 enhancer in Cp also binds another cellular factor, C promoter binding factor 2 (CBF2), whose protein product(s) has not yet been identified. Within the EBNA2 enhancer in Cp, we have previously identified the DNA sequence required for CBF2 binding and also determined that this element is required for efficient activation of Cp by EBNA2. In this study, the CBF2 activity was biochemically purified and microsequenced. The peptides sequenced were identical to the hnRNP protein AUF1. Antibodies against AUF1 but not antibodies to related hnRNP proteins reacted with CBF2 in gel mobility shift assays. In addition, stimulation of the cellular cyclic AMP (cAMP)/protein kinase A (PKA) signal transduction pathway results in an increase in detectable CBF2/AUF1 binding activity extracted from stimulated cells. Furthermore, the CBF2 binding site was able to confer EBNA2 responsiveness to a heterologous promoter when transfected cells were treated with compounds that activate PKA or by cotransfection of plasmids expressing a constitutively active catalytic subunit of PKA, EBNA2-mediated stimulation of the latency Cp is also increased in similar cotransfection assays. These results further support an important role for CBF2 in mediating EBNA2 transactivation; they identify the hnRNP protein AUF1 as a major component of CBF2 and are also the first evidence of a cis-acting sequence other than a CBF1 binding element that is able to confer responsiveness to EBNA2.