Interactions between Egr1 and AP1 factors in regulation of tyrosine hydroxylase transcription
Interactions between Egr1 and AP1 factors in regulation of tyrosine hydroxylase transcription
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DOI：
10.1016/s0169-328x(03)00047-0
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发表时间：
2003-04-10
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影响因子：
--
通讯作者：
Sabban, EL
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文献类型：
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作者：
Nakashima, A;Ota, A;Sabban, EL
几种调节酪氨酸羟化酶（TH）转录的处理，如体内应激或细胞培养中的12-O-十四烷酰基佛波醇-13-乙酸酯（TPA），诱导Egr 1和AP 1因子。以前，我们确定了一个功能性的Egr 1基序与大鼠TH启动子中的Sp1位点重叠。它对Egr 1的反应也需要AP 1/Ebox基序的存在。在这里，我们进一步研究了这些网站之间的串扰。在Sp1/Egr 1和AP 1/Ebox元件之间插入10-或20-bp，降低了Egr 1上调PC 12细胞中由大鼠TH启动子近端272个核苷酸控制的荧光素酶报告活性的能力。用TPA处理的细胞的核提取物进行电泳迁移率变动测定，以鉴定结合AP 1/Ebox基序的因子的组成，以及是否存在与结合Sp1/Egr 1基序的因子的竞争。标记的AP 1/E box寡核苷酸与抗血清形成的复合物被还原或超位移。Fos家族、c-Fos、Fra-2和Jun D.过量的Sp1/Egr 1寡核苷酸或抗Egr 1抗血清不竞争。TPA后2-4 h，Fra-2是复合物的主要成分。用Fra-2转染PC 12细胞诱导的报告活性需要AP 1，但不需要Egr 1基序。然而，当与Fra-2共转染时，Egr 1表达质粒引起的荧光素酶活性的诱导比单独使用Egr 1观察到的低。我们的研究结果表明，虽然它不竞争结合的启动子，Egr 1可以调节TH转录的调节AP 1因子。(C)2003 Elsevier Science B. V.保留所有权利。
Several treatments which regulate tyrosine hydroxylase (TH) transcription, such as stress in vivo, or 12-O-tetradecanoylphorbol-13-acetate (TPA) in cell culture, induce both Egr1 and AP1 factors. Previously, we identified a functional Egr1 motif overlapping with Sp1 site in the rat TH promoter. Its response to Egr1 also required the presence of an AP1/Ebox motif. Here, we further examined the cross-talk between these sites. Insertion of 10- or 20-bp between the Sp1/Egr1 and AP1/Ebox elements, reduced the ability of Egr1 to upregulate luciferase reporter activity controlled by the proximal 272 nucleotides of the rat TH promoter in PC12 cells. Electrophoretic mobility shift assays with nuclear extracts from TPA treated cells were used to identify the composition of the factors which bound the AP1/Ebox motif and whether there is competition with factors which bind the Sp1/Egr1 motif. The complexes formed with labeled AP1/E box oligonucleotide were reduced or supershifted with antisera. to Fos family, c-Fos, Fra-2, and Jun D. Excess Sp1/Egr1 oligonucleotide or anti Egr1 antisera did not compete. Fra-2 was a major component of the complex after 2-4 h TPA. Transfection of PC12 cells with Fra-2 induced reporter activity requiring the AP1, but not the Egr1 motif. However, when cotransfected with Fra-2, Egr1 expression plasmids elicited lower induction of luciferase activity than observed with Egr1 alone. Our results suggest that although it does not compete for binding to the promoter, Egr1 can modulate the regulation of TH transcription by AP1 factors. (C) 2003 Elsevier Science B.V. All rights reserved.