Palvimo J J, Partanen M, Jänne O A
Department of Physiology, University of Helsinki, Finland.
Biochem J. 1996 Jun 15;316 ( Pt 3)(Pt 3):993-8. doi: 10.1042/bj3160993.
The promoter of the murine ornithine decarboxylase (ODC) gene contains, adjacent to the TATA box, a cAMP response element (CRE)-like motif that interacts with specific nuclear proteins. Here we examine the role of this CRE-like element (CREL) in ODC promoter activation in proliferating cells. Mutations that abolished binding of nuclear proteins to CREL influenced only marginally the cAMP induction of the reporter constructs driven by 1.6 kb of the ODC promoter. Instead, these mutations altered the basal promoter function in a cell-specific manner, in that they reduced the promoter activity in CV-1 cells, but increased it in NIH/3T3, CHO and HeLa cells. Thus, depending on the cell type, the CREL motif is able to confer either repression or activation on ODC gene transcription. In contrast with 1.6 kb promoter constructs, the same mutations in the context of a shorter sequence (proximal 133 nt) reduced the promoter strength in all cell types studied. The ability of the CREL element to attenuate transcription seems to be connected with the function of some upstream regulatory elements. Differences in nuclear proteins binding to CREL, as studied by electrophoretic mobility shift assays (EMSAs), did not explain the findings on cell-type specificity in transcriptional activation, as mutations in CREL abrogated formation of specific CREL-protein complexes in all cell lines examined. The protein complexes interacting with CREL were not recognized by antibodies specific for CRE-binding proteins CREB-1 and CREB-2, or activating transcription factors ATF-1, ATF-2 and ATF-3. EMSA experiments also demonstrated co-operative interactions between the CREL motif-binding proteins and other nuclear proteins, such as Sp1, interacting with CG-rich sequences of the promoter. In conclusion, the proximal ODC promoter contains a well-conserved regulatory element, which is clearly different from the CRE/ATF element. This motif acts in concert with other distal and proximal elements in a complex cell-specific manner.
小鼠鸟氨酸脱羧酶（ODC）基因的启动子在TATA框附近含有一个与特定核蛋白相互作用的环磷酸腺苷反应元件（CRE）样基序。在此，我们研究了这个CRE样元件（CREL）在增殖细胞中ODC启动子激活过程中的作用。消除核蛋白与CREL结合的突变对由1.6 kb ODC启动子驱动的报告基因构建体的cAMP诱导仅产生轻微影响。相反，这些突变以细胞特异性方式改变了基础启动子功能，因为它们降低了CV-1细胞中的启动子活性，但在NIH/3T3、CHO和HeLa细胞中则增加了启动子活性。因此，根据细胞类型的不同，CREL基序能够对ODC基因转录产生抑制或激活作用。与1.6 kb启动子构建体不同，在较短序列（近端133 nt）背景下的相同突变降低了所有研究细胞类型中的启动子强度。CREL元件减弱转录的能力似乎与一些上游调控元件的功能有关。通过电泳迁移率变动分析（EMSA）研究发现，与CREL结合的核蛋白差异并不能解释转录激活中细胞类型特异性的结果，因为CREL中的突变消除了所有检测细胞系中特异性CREL-蛋白复合物的形成。与CREL相互作用的蛋白复合物不能被CRE结合蛋白CREB-1和CREB-2或激活转录因子ATF-1、ATF-2和ATF-3的特异性抗体识别。EMSA实验还证明了CREL基序结合蛋白与其他核蛋白（如与启动子富含CG序列相互作用的Sp1）之间的协同相互作用。总之，近端ODC启动子含有一个保守的调控元件，它明显不同于CRE/ATF元件。这个基序以复杂的细胞特异性方式与其他远端和近端元件协同作用。
