Mantis N J, Winans S C
Section of Microbiology, Cornell University, Ithaca, New York 14853.
J Bacteriol. 1992 Feb;174(4):1189-96. doi: 10.1128/jb.174.4.1189-1196.1992.
A set of Agrobacterium tumefaciens operons required for pathogenesis is coordinately induced during plant infection by the VirA and VirG proteins. The intracellular concentration of VirG increases in response to acidic media, and this response was proposed to be regulated at the level of transcription at a promoter (P2) that resembles the Escherichia coli heat shock promoters. To test this hypothesis, we first constructed a virG-lacZ transcriptional fusion. A strain containing this fusion had higher levels of beta-galactosidase activity in acidic media than in media at neutral pH. Second, primer extension analysis of virG indicated that acidic media stimulated the transcription of this promoter. To determine whether P2 is a member of a heat shock-like regulon in A. tumefaciens, five agents that induce E. coli heat shock genes were tested for their abilities to induce a P2-lacZ fusion in A. tumefaciens. P2 was most strongly induced by low pH, was moderately stimulated by CdCl2 or mitomycin C, and was slightly induced by P2 as measured by beta-galactosidase activity and primer extension analysis. Induction by these treatments did not require any Ti plasmid-encoded function or the chromosomally encoded RecA protein. We also pulse-labeled cellular proteins after a shift to low pH and detected several proteins whose synthesis was induced by these conditions. We conclude that P2 is primarily induced by acid pH and secondarily by certain other stimuli, each of which is stressful to cell growth. This stress induction is at least partly independent of the heat shock and SOS responses.
一组参与致病过程的根癌土壤杆菌操纵子在植物感染期间由VirA和VirG蛋白协同诱导。VirG的细胞内浓度会因酸性培养基而增加,并且有人提出这种反应是在一个类似于大肠杆菌热休克启动子的启动子(P2)的转录水平上受到调控。为了验证这一假设,我们首先构建了一个virG-lacZ转录融合体。含有这种融合体的菌株在酸性培养基中的β-半乳糖苷酶活性水平高于中性pH培养基中的水平。其次,对virG进行引物延伸分析表明,酸性培养基刺激了该启动子的转录。为了确定P2是否是根癌土壤杆菌中热休克样调控子的一员,测试了五种诱导大肠杆菌热休克基因的试剂在根癌土壤杆菌中诱导P2-lacZ融合体的能力。通过β-半乳糖苷酶活性和引物延伸分析测量,P2在低pH条件下诱导最强,在CdCl2或丝裂霉素C作用下受到中度刺激,在P2作用下受到轻微诱导。这些处理的诱导不需要任何Ti质粒编码的功能或染色体编码的RecA蛋白。我们还在转移到低pH后对细胞蛋白进行脉冲标记,并检测到几种在这些条件下合成被诱导的蛋白。我们得出结论,P2主要由酸性pH诱导,其次由某些其他刺激诱导,每种刺激对细胞生长都有压力。这种应激诱导至少部分独立于热休克和SOS反应。