Hong G F, Burn J E, Johnston A W
AFRC Institute of Plant Science Research, John Innes Institute, Norwich, UK.
Nucleic Acids Res. 1987 Dec 10;15(23):9677-90. doi: 10.1093/nar/15.23.9677.
In Rhizobium leguminosarum biovar viciae, the regulatory nodulation nodD gene has at least two functions. It constitutively represses its own transcription and in the presence of inducer flavonoid molecules, it activates the expression of two other nod gene transcriptional units, nodABCIJ and nodFE. Upstream of nodA and nodF is a conserved sequence, the nod box, which has been implicated in nodD-mediated transcriptional activation of these genes. DNA fragments spanning the nod boxes that precede nodA and nodF were end-labelled and were exposed to cell-free extracts obtained from strains of Rhizobium. Using the gel retardation technique, it was shown that a complex between protein and these DNA fragments was formed, but only if the extract contained a functional nodD gene. Evidence that the protein that binds to the regulatory sequences is the nodD gene product came from the observation that a complex was formed between the nod box preceding nodA and protein from a cell-free extract isolated from Escherichia coli containing the cloned nodD gene. Extracts from Rhizobium strains containing mutant forms of nodD which were specifically affected in autoregulation or in flavonoid-dependent activation formed either no protein DNA complex or formed a complex with altered mobility compared to that obtained with extracts from wild-type strains.
在豌豆根瘤菌蚕豆生物变种中，调控结瘤的nodD基因至少有两种功能。它组成型地抑制自身转录，并且在诱导剂类黄酮分子存在的情况下，它激活另外两个nod基因转录单元nodABCIJ和nodFE的表达。nodA和nodF的上游是一个保守序列，即nod框，它与nodD介导的这些基因的转录激活有关。跨越nodA和nodF之前的nod框的DNA片段进行了末端标记，并与从根瘤菌菌株中获得的无细胞提取物一起孵育。使用凝胶阻滞技术表明，只有当提取物含有功能性nodD基因时，才会形成蛋白质与这些DNA片段之间的复合物。与nodD基因产物结合的蛋白质的证据来自以下观察结果：在nodA之前的nod框与来自含有克隆的nodD基因的大肠杆菌分离的无细胞提取物中的蛋白质之间形成了复合物。含有在自调控或类黄酮依赖性激活中受到特异性影响的nodD突变形式的根瘤菌菌株的提取物，要么不形成蛋白质-DNA复合物，要么形成与野生型菌株提取物相比迁移率改变的复合物。
