Interactions between c-Jun, nuclear factor 1, and JC virus promoter sequences: Implications for viral tropism
Interactions between c-Jun, nuclear factor 1, and JC virus promoter sequences: Implications for viral tropism
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DOI：
10.1128/jvi.01355-06
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发表时间：
2006-11-01
影响因子：
5.4
通讯作者：
Major, Eugene O.
中科院分区：
文献类型：
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作者：
Ravichandran, Veerasarny;Sabath, Bruce F.;Major, Eugene O.
人多核病毒 JC (JCV) 的感染周期最终在细胞核中的病毒蛋白表达和基因组复制水平上受到调节。这种活性是由 JCV 调节区（启动子）内的变异核苷酸序列与结合特定 DNA 共有位点的细胞转录因子之间的相互作用促进的。在之前的工作中，我们确定了 NF-1 类成员 NF-1X 是影响 JCV 细胞宿主范围的关键转录因子。在变体 JCV 启动子以及其他病毒和细胞启动子中，经常会发现相邻的 NF-1 和 AP-1 共有位点。这两个结合位点非常接近，表明 NF-1 和 AP-1 蛋白之间存在相互作用的机会。在这里，通过电泳迁移率变动分析，我们显示了 AP-1 家族成员 c-Jun 对 NF-1 蛋白结合源自 JCV 启动子序列的 NF-1 共有位点的时间和剂量依赖性干扰。此外，正如蛋白质-蛋白质相互作用测定所证明的，我们确定了 NF-1X 和 c-Jun 之间独立于 DNA 结合的特异性结合亲和力。最后，为了比较 NF-1X 和 c-Jun 在 JCV 启动子序列上的结合特征，同时体内检测病毒活性水平，我们开发了一种锚定转录启动子 (ATP) 测定法。使用转染过表达 NF-1X 或 c-Jun 的 JCV 感染细胞的提取物，ATP 测定显示 NF-1X 结合和病毒蛋白表达同时增加。相反，c-Jun 结合的增加伴随着 NF-1X 结合和病毒蛋白表达的减少。因此，c-Jun 对 NF-1X 结合的抑制似乎在调节 JCV 活性水平中发挥作用。
The infectious cycle of the human polyornavirus JC (JCV) is ultimately regulated in cellular nuclei at the level of viral protein expression and genomic replication. Such activity is prompted by interactions between variant nucleotide sequences within the JCV regulatory region (promoter) and cellular transcription factors that bind specific DNA consensus sites. In previous work we identified an NF-1 class member, NF-1X, as a critical transcription factor affecting the JCV cellular host range. Within variant JCV promoters, as well as other viral and cellular promoters, adjacently located NF-1 and AP-1 consensus sites are often found. The close proximity of these two binding sites suggests the opportunity for interaction between NF-1 and AP-1 proteins. Here, by electrophoretic mobility shift assays, we show temporal and dose-dependent interference by an AP-1 family member, c-Jun, upon NF-1 proteins binding an NF-1 consensus site derived from JCV promoter sequence. Moreover, as demonstrated by protein-protein interaction assays, we identify specific binding affinity independent of DNA binding between NF-1X and c-Jun. Finally, to compare the binding profiles of NF-1X and c-Jun on JCV promoter sequence in parallel with in vivo detection of viral activity levels, we developed an anchored transcriptional promoter (ATP) assay. With use of extracts from JCV-infected cells transfected to overexpress either NF-1X or c-Jun, ATP assays showed concurrent increases in NF-1X binding and viral protein expression. Conversely, increased c-Jun binding accompanied decreases in both NF-1X binding and viral protein expression. Therefore, inhibition of NF-1X binding by c-Jun appears to play a role in regulating levels of JCV activity.