RNA interference and low-allergenic plant-derived food under changing environmental conditions

番茄（Solanum esculentum）引起过敏症状，如口腔过敏综合征、接触性荨麻疹综合征或过敏患者的过敏性休克。到目前为止，至少有13种蛋白质与番茄中的ige结合有关。这些ige结合蛋白中有四种被归类为过敏原。在过去的几年里，我们建立了番茄果实作为模型来测试产生低过敏性食物的分子策略。这个子项目的主要目的是(i)提供原理证明，即使需要靶向多个过敏原，并且涉及到具有基本细胞功能的过敏原，RNAi技术也可以用于产生低过敏性食物，（ii）探索靶向基因组修饰的新技术，（iii）通过酵母随机诱变结合功能选择来研究profilin的致敏结构。为了实现我们的第一个目标，我们已经开发了一种育种方案，将番茄中多达8种ige结合蛋白的下调结合起来。第二个目标将通过使用定制的TALE核酸酶诱变lyce3来实现。为了随机确定谱蛋白的致敏决定因素，我们开发了一个基于酵母的筛选平台，以选择功能性和低致敏性的谱蛋白变体。该系统已成功用于鉴定低过敏性Lyc e1，并将用于筛选来自其他植物来源的低过敏性谱变异，如Bet v 2或Api g 4。在反向实验中，我们将诱变酵母profilin，以测试是否有可能将非过敏原蛋白转化为过敏原。

Tomato (Solanum esculentum) elicits allergic symptoms such as oral allergy syndrome, contact urticarial syndrome or anaphylactic shock in sensitized patients. So far at least 13 proteins have been associated with IgE-binding in tomato. Four of these IgE-binding proteins have been classified as allergens. Over the last years we have established tomato fruits as models to test molecular strategies to generate hypoallergenic food. Major aims of this sub-project are (i) to provide proof of principle that RNAi technologies can be used to generate hypoallergenic food even if multiple allergens need to be targeted and if allergens with essential cellular functions are involved, (ii) to explore new technologies for targeted genome modifications and (iii) to investigate allergenic structures of profilin by random mutagenesis combined with functional selection in yeast. To achieve our first aim we have developed a breeding scheme to combine down-regulation of up to eight IgE-binding proteins from tomato. The second goal will be reached by using customized TALE nucleases for mutagenesis of Lyc e 3. To identify allergenic determinants of profilins in a random fashion we have developed a yeast-based screening platform to allow selection for functional and hypoallergenic profilin variants. This system has successfully been used to identify hypoallergenic Lyc e 1 and will be applied to screen for hypoallergenic profilin variants from other plant sources such as Bet v 2 or Api g 4. In a reverse experiment we will mutagenize yeast profilin to test whether it is possibly to convert a non-allergenic protein into an allergen.