Molecular Genetics of Pectin Lyase Regulation in Erwinia

欧文氏菌果胶裂解酶调节的分子遗传学

• 批准号: 9018733
• 负责人: Arun Chatterjee
• 金额: $ 28.5万
• 依托单位: University of Missouri-Columbia
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-02-01 至 1994-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9018733&HistoricalAwards=false
• 关键词: Molecular; Genetics; Pectin; Lyase; Regulation

Erwinia carotovora subsp. carotovora (hereafter Ecc) and most other soft-rot Erwinia spp. produce an assortment of extracellular degradative enzymes such as pectin lyase (Pnl), pectate lyase (Pel), polygalacturonase (Peh), cellulase, protease and phospholipase. A long-term goal of our research is to clarify the genetic and physiological aspects of extracellular enzyme production in soft-rot Erwinia and to elucidate the role of these enzymes in bacterial plant pathogenicity. We have shown that while pectinolytic (Pnl) and pectolytic (Pel and Peh) enzymes cause maceration of plant tissue, they are regulated differently. In contrast to pectolytic enzymes, most of which are induced by catabolic products of polygalacturonate or pectin, Pnl is induced by DNA-damaging agents such as mitomycin C, nalidixic acid, or ultraviolet light. In Ecc strain 71 (Ecc71) the induction of pnlA, the pectin lyase structural gene, by these DNA-damaging agents requires products of recA and an activator gene, digR (= regulator of damage-inducible genes). These products also appear to control bacteriocin (carotovoricin, Ctv) production and cellular lysis in this bacterium. We have reconstituted and inducible Pnl system in a RecA+ E. coli strain carrying a pnlA+ plasmid and a digR+ plasmid. The E. coli strain responds to the same regulatory signals as the Ecc71 Pnl producing system. Moreover, our recent findings indicate that digR and not pnlA is induced by DNA damaging agents in a RecA+ LexA+ E. coli. These observations suggest a cascade type of regulation wherein a regulatory gene (digR), required for the transcription of pnlA and possibly other damage-inducible genes, is controlled by DNA damaging agents. The objectives of this research are (a) to identify and characterize upstream DNA segments of pnlA that regulate transcription, (b) to characterize the activator gene, digR and to identify its product, (c) to elucidate the expression of digR, and (d) to understand the mechanism by which DigR activates pnlA transcription. In these studies we intend to utilize genetic and molecular technologies and various tools already available in E. coli and in Ecc71. The emerging knowledge should help in defining the regulatory circuit and in understanding the molecular events in the expression of a plant virulence factor by DNA-damaging agents.

胡萝卜软腐欧文氏菌胡萝卜素（以下简称Ecc）和大多数 其他软腐欧文氏菌属生产各种各样的 胞外降解酶如果胶裂解酶（Pnl）， 果胶酸裂解酶（Pel），多聚半乳糖醛酸酶（Peh），纤维素酶，蛋白酶 和磷脂酶。 我们研究的长期目标是 阐明细胞外遗传和生理方面 软腐欧文氏菌酶的生产，并阐明作用 这些酶在细菌植物致病性中的作用。 我们已经表明 虽然果胶溶解的（Pnl）和果胶溶解的（Pel和Peh） 酶引起植物组织的浸渍，它们被调节 不同. 与大多数果胶酶相反， 是由聚半乳糖醛酸或果胶的分解代谢产物诱导的， Pn1由DNA损伤剂如丝裂霉素C诱导， 萘二甲酸或紫外线。 在Ecc菌株71（Ecc 71）中 果胶裂解酶结构基因pnlA的诱导， DNA损伤剂需要recA和激活剂的产物 损伤诱导基因的调节子。 这些 产品似乎也控制细菌素（胡萝卜素，ctv） 生产和细胞裂解。 我们有 在RecA + E中重构和诱导的Pnl系统。杆菌菌株 携带pnlA+质粒和digR+质粒。 急诊杆菌菌株 响应与Ecc 71 Pnl相同的调节信号 生产系统。 此外，我们最近的研究结果表明， 在RecA+细胞中，digR而不是pnlA被DNA损伤剂诱导。 莱克萨+E.杆菌 这些观察结果表明， 调节，其中调节基因（digR），所需的 pnlA和可能的其它损伤诱导基因的转录， 是由DNA破坏剂控制的 这一目标 研究是（a）识别和表征上游DNA 调节转录的pnlA片段，（B）表征 激活基因digR并鉴定其产物，（c） 阐明digR的表达，和（d）了解 DigR激活pnlA转录的机制。 在这些 我们打算利用基因和分子技术进行研究， 以及E.大肠杆菌Ecc71. 的 新出现的知识应该有助于确定调节回路 以及了解基因表达中的分子事件 植物毒力因子的DNA损伤剂。