Ramachandran A, Jain A, Arora P, Bashyam M D, Chatterjee U, Ghosh S, Parnaik V K, Hasnain S E
Eukaryotic Gene Expression Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
J Biol Chem. 2001 Jun 29;276(26):23440-9. doi: 10.1074/jbc.M101537200. Epub 2001 Apr 6.
We earlier documented the involvement of a cellular factor, polyhedrin (polh) promoter-binding protein, in transcription from the Autographa californica nuclear polyhedrosis virus polh gene promoter. Sequences upstream of the polh promoter were found to influence polh promoter-driven transcription. Analysis of one such region, which could partially compensate for the mutated polh promoter and also activate transcription from the wild-type promoter, revealed a sequence (AcSp) containing a CACCC motif and a loose GC box resembling the binding motifs of the transcription factor Sp1. AcSp and the consensus Sp1 sequence (cSp) specifically bound factor(s) in HeLa and Spodoptera frugiperda (Sf9) insect cell nuclear extracts to generate identical binding patterns, indicating the similar nature of the factor(s) interacting with these sequences. The AcSp and cSp oligonucleotides enhanced in vivo expression of a polh promoter-driven luciferase gene. In vivo mopping of these factor(s) significantly reduced transcription from the polh promoter. Recombinant viruses carrying deletions in the upstream AcSp sequence confirmed the requirement of these factor(s) in polh promoter-driven transcription in the viral context. We demonstrate for the first time DNA-protein interactions involving novel members of the Sp family of proteins in adult insect cells and their involvement in transcription from the polh promoter.
我们之前记录了一种细胞因子,即多角体蛋白(polh)启动子结合蛋白,参与了苜蓿银纹夜蛾核型多角体病毒polh基因启动子的转录过程。研究发现,polh启动子上游的序列会影响polh启动子驱动的转录。对其中一个区域进行分析,该区域能够部分补偿突变的polh启动子并激活野生型启动子的转录,结果揭示了一个包含CACCC基序和类似转录因子Sp1结合基序的松散GC框的序列(AcSp)。AcSp和共有Sp1序列(cSp)在HeLa和草地贪夜蛾(Sf9)昆虫细胞核提取物中特异性结合因子,产生相同的结合模式,表明与这些序列相互作用的因子具有相似的性质。AcSp和cSp寡核苷酸增强了polh启动子驱动的荧光素酶基因的体内表达。体内清除这些因子显著降低了polh启动子的转录。携带上游AcSp序列缺失的重组病毒证实了在病毒环境中polh启动子驱动的转录需要这些因子。我们首次证明了在成年昆虫细胞中涉及Sp蛋白家族新成员的DNA-蛋白质相互作用及其参与polh启动子的转录过程。