GALLS, A Novel Agrobacterium Rhizogenes Protein Required for Gene Transfer to Plants

农杆菌和根状芽孢杆菌将基因和蛋白质转移到植物细胞中。转移的DNA在植物细胞中稳定地遗传和表达，引起冠瘿病或毛状根病。这是已知的细菌和高等生物之间自然基因交换的唯一例子。植物分子生物学家创造了“解除武装”的瘤胃杆菌菌株，这种菌株缺乏致病基因，但保留了将DNA转移到植物中所需的其他基因。农杆菌是向植物传递基因的首选方法。这项技术被广泛用于培育转基因作物，2003年在1.06亿英亩的土地上种植了转基因作物，为美国农民节省了18亿美元，同时减少了4580万磅农药的使用。农杆菌将DNA转移到植物细胞核的能力取决于DNA结合蛋白（连同DNA）从细菌细胞转移到植物细胞。一些根状芽孢杆菌缺乏这种关键的蛋白质，但它们仍然能够将DNA转移到植物细胞核中。一种不寻常的根状芽孢杆菌蛋白（GALLS）为植物细胞核吸收DNA提供了另一种途径。GALLS蛋白位于细胞核内，与另一种蛋白相互作用，后者附着在转移的DNA上。GALLS蛋白似乎将进入的细菌DNA拉入植物细胞核。为了验证这一假设，将测试GALLS蛋白是否具有将DNA动员到细胞核所需的预测生化活性。几种遗传和生化方法将用于鉴定与细胞核中GALLS蛋白相互作用的植物蛋白。细胞生物学和荧光显微镜将用于监测GALLS蛋白从细菌细胞到植物细胞的运输。这项工作可能通过将传入的细菌DNA靶向到宿主植物DNA的特定位置来改进植物转化技术。因此，新基因可以引入到不破坏重要植物基因的位置，从而避免发育异常，这是目前植物转化方法中常见的。更广泛的影响。本科生、高中生和研究生将接受密集的研究训练。作为推广活动，PI将在夏季植物生物技术讲习班上培训高中教师。

Agrobacterium tumefaciens and A. rhizogenes transfer genes and proteins into plant cells. Transferred DNA is stably inherited and expressed in plant cells, causing crown gall or hairy root disease. These are the only known examples of natural genetic exchange between bacteria and higher organisms. Plant molecular biologists created "disarmed" strains of A. tumefaciens that lack the disease-causing genes but retain other genes needed to transfer DNA into plants. Agrobacterium is the preferred method to deliver genes to plants. This technology is widely used to create transgenic crop plants, which were planted on 106 million acres in 2003 and saved US farmers $1.8 billion while reducing pesticide use by 45.8 million pounds. The ability of A. tumefaciens to transfer DNA into the plant cell nucleus depends on a DNA-binding protein that is transported (along with DNA) from bacterial cells into plant cells. Some strains of A. rhizogenes lack this crucial protein, but they are still able to transfer DNA into the plant cell nucleus. An unusual A. rhizogenes protein (GALLS) provides an alternative route for uptake of DNA into the plant cell nucleus. The GALLS protein is located inside the nucleus where it interacts with another protein, which is attached to the transferred DNA. The GALLS protein appears to pull the incoming bacterial DNA into the nucleus of the plant cell. To test this hypothesis, GALLS protein will be tested for the predicted biochemical activity needed to mobilize DNA into the nucleus. Several genetic and biochemical approaches will be used to identify plant proteins that interact with GALLS protein in the nucleus. Cell biology and fluorescence microscopy will be used to monitor transport of GALLS protein from bacterial cells into plant cells. This work may improve plant transformation technology by making it possible to target incoming bacterial DNA into specific locations in the host plant's DNA. Thus, new genes can be introduced into locations that do not disrupt important plant genes, thereby avoiding developmental abnormalities, which are common using current plant transformation methods. Broader Impacts. Undergraduates, high school students, and graduate students will receive intensive research training. As outreach, the PI will train high school teachers in a summer workshop on plant biotechnology.