Mao C, Downie J A, Hong G
Shanghai Institute of Biochemistry, China.
Gene. 1994 Jul 22;145(1):87-90. doi: 10.1016/0378-1119(94)90327-1.
In Rhizobium leguminosarum (R.l.) biovar viciae, the nodulation gene nodD encodes a transcriptional activator (NodD) which binds to highly conserved DNA sequences (nod-boxes) in the promoters of other nod operons. In addition, NodD represses nodD transcription and this occurs at the divergent and overlapping nodA-nodD promoters. We mutagenised this region with hydroxylamine, and by cloning the mutagenised DNA into a vector carrying the lacZ reporter gene downstream from the cloning site identified mutations affecting nodD expression and repression. The resulting plasmids were transferred to R. l. viciae strains containing or lacking nodD. Two classes of promoter mutants were identified: those in which nodD transcription was altered and those in which NodD-dependent repression was altered. The nucleotide (nt) changes in the promoter region were found to be located within two inverted repeat sequences (A2 and A3) which are about 70 bp apart. A2 is important for nodD transcription and A3 (which is upstream from A2) is involved in NodD-dependent repression. The nt sequence at A3 shows some homology to the nod-box region of the nodA promoter. It is proposed that the NodD-dependent repression occurs as a result of NodD binding to both A3 and the nodA nod-box, forming a loop which prevents transcription of nodD from its promoter, A2, which lies between A3 and the nod-box. This model is supported by the observation that there are at least three sites for NodD binding in the nodA-nodD promoter region.
在豌豆根瘤菌（R.l.）蚕豆生物变种中，结瘤基因nodD编码一种转录激活因子（NodD），它能与其他结瘤操纵子启动子中高度保守的DNA序列（结瘤框）结合。此外，NodD会抑制nodD转录，这种抑制发生在反向且重叠的nodA - nodD启动子处。我们用羟胺诱变了该区域，并通过将诱变后的DNA克隆到一个在克隆位点下游携带lacZ报告基因的载体中，鉴定出影响nodD表达和抑制的突变。将所得质粒转移到含有或缺乏nodD的R.l.蚕豆菌株中。鉴定出了两类启动子突变体：一类是nodD转录发生改变的突变体，另一类是NodD依赖性抑制发生改变的突变体。发现启动子区域的核苷酸（nt）变化位于两个相距约70 bp的反向重复序列（A2和A3）内。A2对nodD转录很重要，而A3（位于A2上游）参与NodD依赖性抑制。A3处的nt序列与nodA启动子的结瘤框区域有一些同源性。有人提出，NodD依赖性抑制是由于NodD与A3和nodA结瘤框都结合，形成一个环，从而阻止nodD从位于A3和结瘤框之间的其启动子A2转录。这一模型得到了以下观察结果的支持：在nodA - nodD启动子区域至少有三个NodD结合位点。
