Establishment of a library of membrane and effector proteins of ´Candidatus P. mali´ to identify host-(apple)-pathogen interactions and to disrupt them

候选植物：马里植原体马利杆菌是一种引起苹果增殖的植物病原韧皮部定殖原核生物，是欧洲所有温和气候产区苹果的重要经济病害。该疾病是由属于仙人掌属的昆虫媒介传播的。到目前为止，苹果增殖病的防治还只能靠施用杀虫剂。直到最近，植物原体在无细胞培养基中抵抗了所有的培养尝试，因此这些病原体在生理和生化基础上的特征很差。最近，研究人员发现了Ca。对马里疟原虫和其他三种植物浆体基因组进行了完全测序。该基因组约为60万bp，有497个开放阅读框。马里疟原虫在大小和组织结构上不同于其他三种已知的植原体基因组。初步结果表明，免疫优势膜蛋白和植物原体效应蛋白都与植物原体疾病的发病机制有关。因此，我们将建立酵母双杂交（Y2H）酵母膜蛋白和Ca效应蛋白文库。利用来自Malus x domestica cv Golden（“诱饵”）cDNA文库的Y2H文库筛选宿主（苹果）蛋白与植物原体蛋白的相互作用。这些相互作用蛋白的鉴定将有助于更好地了解苹果增殖病的发病机制，并有助于制定创新的管理策略。因此，我们将通过噬菌体展示选择能够干扰和/或破坏这些相互作用的适体。此外，我们将评估这些适体的应用策略。

Candidatus Phytoplasma mali (´Ca. P.mali´) is a plant pathogenic phloem colonizing prokaryote causing apple proliferation, an economically important disease in apple in all European production areas with moderate climates. The disease is transmitted by insect vectors belonging to the genus Cacopsylla. Up to now the apple proliferation disease is controlled only by the application of insecticides. Until recently phytoplasmas have resisted all attempts of cultivation in cell free media, therefore these pathogens are poorly characterized on a physiological and biochemical basis. Recently the genome of ´Ca. P. mali´ and three other phytoplasmas genomes were completely sequenced. With a size of approximately 600.000 bp and 497 open reading frames the genome of `Ca. P. mali´ differs in size and organisation from the three other known phytoplasma genomes.First results indicate that both immunodominant membrane proteins and putative effector proteins of phytoplasmas are relevant in the pathogenesis of phytoplasma diseases. Therefore we will establish a yeast two hybrid (Y2H) library of membrane and effector proteins of ´Ca. P. mali´ ("Prey") to identify interactions of host-(apple-) proteins with phytoplasma proteins based on a screening with a Y2H library deriving from a cDNA library of Malus x domestica cv Golden ("Bait") .The identification of these interacting proteins will lead to a better understanding of the pathogenesis of the apple proliferation disease and moreover, enable the development of innovative management strategies. In consequence we will select aptamers via phage display which are able to disturb and/or disrupt these interactions. Moreover we will evaluate strategies for the application of these aptamers.