Chou A Y, Archdeacon J, Kado C I
Davis Crown Gall Group, University of California, Davis, CA 95616, USA.
Proc Natl Acad Sci U S A. 1998 Apr 28;95(9):5293-8. doi: 10.1073/pnas.95.9.5293.
Virulence genes of Agrobacterium tumefaciens are under the control of positive and negative transcriptional regulators. We found that the transcriptional regulator Ros controls expression of the plant oncogene ipt, which encodes isopentenyl transferase, in A. tumefaciens. This enzyme is involved in biosynthesis of the plant growth hormone cytokinin in the host plant. An ipt promoter::cat reporter gene fusion showed a 10-fold increase in ipt promoter activity in A. tumefaciens ros mutant strains when compared with wild type. Also, increased levels (10- to 20-fold) of isopentenyl adenosine, the product of the reaction catalyzed by isopentenyl transferase, were detected in ros mutant strains. In vitro studies using purified Ros showed it binds directly to the ipt promoter. Analysis of the deduced Ros amino acid sequence identified a novel type of C2H2 zinc finger. In Ros the peptide loop spacing of the zinc finger is 9 amino acids as opposed to the invariant 12 amino acids in the classical C2H2 motif. Site-directed mutagenesis of Cys-82 and His-92 in this motif showed that these residues are essential for Zn2+ and DNA binding activities of Ros. The existence of such a regulator in Agrobacterium may be due to horizontal interkingdom retrotransfer of the ros gene from plant to bacteria.
根癌土壤杆菌的毒力基因受正、负转录调节因子的控制。我们发现转录调节因子Ros控制根癌土壤杆菌中植物致癌基因ipt的表达，ipt编码异戊烯基转移酶。这种酶参与宿主植物中植物生长激素细胞分裂素的生物合成。与野生型相比，ipt启动子::cat报告基因融合体在根癌土壤杆菌ros突变株中显示出ipt启动子活性增加了10倍。此外，在ros突变株中检测到异戊烯基转移酶催化反应产物异戊烯基腺苷的水平增加（10至20倍）。使用纯化的Ros进行的体外研究表明它直接与ipt启动子结合。对推导的Ros氨基酸序列的分析鉴定出一种新型的C2H2锌指。在Ros中，锌指的肽环间距为9个氨基酸，而经典C2H2基序中不变的是12个氨基酸。对该基序中Cys-82和His-92进行定点诱变表明，这些残基对于Ros的Zn2+和DNA结合活性至关重要。根癌土壤杆菌中存在这样一种调节因子可能是由于ros基因从植物到细菌的水平跨王国逆转录转移所致。
