Lauridsen P, Franssen H, Stougaard J, Bisseling T, Marcker K A
Department of Molecular Biology, University of Aarhus, Denmark.
Plant J. 1993 Mar;3(3):483-92. doi: 10.1111/j.1365-313x.1993.tb00168.x.
The beta-glucuronidase (GUS) activity expressed from the soybean early nodulin ENOD2(B) gene promoter was localized histochemically in nodules of Lotus corniculatus and Trifolium repens. In both the determinate Lotus nodules and the indeterminate Trifolium nodules, activity was found in the parenchyma cells and especially in cells close to the vascular tissue of nodules. The characteristic cell-specific expression of the soybean ENOD2 gene was therefore maintained by the ENOD2(B) promoter in the two developmentally different nodule types. Important DNA elements recognized in transgenic nodules were identified by deletion and hybrid promoter analysis in Lotus corniculatus. An indispensable positive element (PE) and a possible tissue specific element was defined between positions -1792 and -1582 from the transcription start site. Another qualitative control element located between -380 and -53 conferred the ENOD2 characteristic cell type expression on hybrid promoters. This element contains the conserved nodulin gene sequences CTCTT and AAAGAT. In contrast to the ENOD2(B) promoter a chimeric leghemoglobin Ibc3-GUS gene was expressed in the infected cells of both types of nodules. In the indeterminate nodules expression was restricted to the interzone II-III and the active nitrogen-fixing zone III. Interchange of the distal strong positive element (SPE) of Ibc3 and the ENOD2 positive element resulted in an expression pattern different from that observed for the Ibc3 and ENOD2 genes, indicating that different interactions of trans-acting factors are required for regulation of early as well as late nodulin genes.
大豆早期根瘤素ENOD2(B)基因启动子所表达的β-葡萄糖醛酸酶（GUS）活性通过组织化学方法定位在百脉根和白三叶草的根瘤中。在定型的百脉根根瘤和不定型的白三叶草根瘤中，均在薄壁细胞中发现了活性，尤其是在靠近根瘤维管组织的细胞中。因此，大豆ENOD2基因特有的细胞特异性表达通过ENOD2(B)启动子在两种发育不同的根瘤类型中得以维持。通过对百脉根转基因根瘤进行缺失和杂交启动子分析，鉴定出了在转基因根瘤中识别的重要DNA元件。在转录起始位点上游-1792至-1582位之间确定了一个不可或缺的正向元件（PE）和一个可能的组织特异性元件。位于-380至-53位之间的另一个定性控制元件赋予杂交启动子ENOD2特征性的细胞类型表达。该元件包含保守的根瘤素基因序列CTCTT和AAAGAT。与ENOD2(B)启动子不同，嵌合的豆血红蛋白Ibc3-GUS基因在两种类型根瘤的感染细胞中均有表达。在不定型根瘤中，表达局限于II-III区和活跃的固氮III区。Ibc3的远端强正向元件（SPE）与ENOD2正向元件互换后，导致表达模式不同于Ibc3和ENOD2基因所观察到的模式，这表明早期和晚期根瘤素基因的调控需要反式作用因子的不同相互作用。
