Symbiotic Association of Aphids: Biochemical Role of the Prokaryotic Endosymbiont

9402813鲍曼蚜虫的生存依赖于与原核内共生体的结合，该内共生体位于蚜虫体腔中的特化细胞内。 使用蚜虫Schizaphis graminum（绿芽），我们发现其内共生体（Buchnera aphidicola）具有色氨酸生物合成途径的所有基因，并且trpEG（邻氨基苯甲酸合成酶的基因，该途径的第一种酶）在质粒上扩增。这些结果与过去的观察结果一致，表明内共生体合成色氨酸，这是利用蚜虫宿主。 初步研究结果表明，在远缘内共生体中的Schlectendalia chinensis，tryEG有一个染色体定位。 我们打算继续这项研究，通过检查色氨酸生物合成途径的基因组织在这种蚜虫物种的内共生体，并进行调查，以确定染色体定位的trpEG在其他蚜虫物种的内共生体的基因扩增的分布。 起始芳香族氨基酸（色氨酸、苯丙氨酸、酪氨酸）以及叶酸的生物合成的第一反应由3-脱氧-D-阿拉伯庚酮糖酸-7-磷酸（DAHP）合酶催化。 我们打算确定该基因是否位于质粒上、染色体上或是否存在同功能DAHP酶。 这将通过限制性酶和Southern印迹分析以及酶基因的克隆和测序来实现。 并与E.大肠杆菌和其他微生物。 在大多数原核生物中，核糖体RNA的基因以16 S和5S rRNA的顺序排列为单个转录单位。 我们证明了B. 蚜虫只有一个拷贝的16 S 23 s rRNA基因，并且它与其他rRNA基因不连锁。 我们将通过检查B中23 S和5S rRNA基因的组织来继续这项研究。蚜虫。 将对基因进行克隆、测序，并将上游和下游区域与蚜虫B. 16 S rRNA转录单位的推定启动子和终止子进行比较。蚜虫在其最活跃的生长阶段通过单性生殖产生活的幼仔。 关于蚜虫生长周期内内共生体的数量，目前还不清楚。 采用定量聚合酶链反应，我们将确定B中16 S和23 S rRNA基因的拷贝数。在蚜虫生长周期中的蚜虫-蚜虫DNA制备。 结果将是内共生体数量的指示。 利用免疫学方法，我们将确定几种必需的内共生体成分的量。 这包括核糖体（核糖体蛋白S1），RNA聚合酶（β亚基），σ因子和GroEL。 从这些研究中获得的结果可能是用于最终控制蚜虫种群的操纵的内共生体，这是必不可少的生存。 ***

9402813 Baumann Aphids are dependent for survival on an association with a prokaryotic endosymbiont housed within specialized cells in the aphid body cavity. Using the aphid Schizaphis graminum ( greenbud) we have found that its endosymbiont (Buchnera aphidicola) has all of the genes of the tryptophan biosynthetic pathway and the trpEG (genes for anthranilate synthase, the first enzyme of the pathway) is amplified on a plasmid. These results are consistent with past observations indicating that the endosymbiont synthesizes tryptophan which is utilized by the aphid host. Preliminary results have indicated that, in the distantly related endosymbiont of the aphid Schlectendalia chinensis, tryEG has a chromosomal location. We intend to continue this research by examining the gene organization of the tryptophan biosynthetic pathway in the endosymbiont of this aphid species and making a survey to determine the distribution of gene amplification of chromosomal location of trpEG in the endosymbionts of other aphid species. The first reaction which initiates the biosynthesis of aromatic amino acids (tryptophan, phenylalanine, tyrosine) as well as folic acid is catalyzed by 3-deoxy-D-arabino-heptulosonate-7-phosphate (DAHP) synthase. We intend to determine if this gene is located on a plasmid, on the chromosome or if there are isofunctional DAHP synthases. This will be accomplished by restriction enzyme and Southern blot analysis as well as the cloning and sequencing of the gene(s) for the enzyme(s). Comparisons will be made with the gene organization in E. coli and other organisms. In most prokaryotes the genes for ribosomal RNA are arranged as a single transcription unit in the order 16S, and 5S rRNAs. We have shown that B. aphidicola has only one copy of the 16S 23s rRNA gene and is unusual in that it is unlinked to the other rRNA genes. We will continue this study by examining the organization of the 23S and 5S rRNA genes in B. aphidicola. The genes will be cloned, sequenced, and the up stream and downstream regions compared to the putative promoter and terminator of the B.aphidicola 16S rRNA transcription unit. In their most active stage of growth aphids reproduce by parthenogenesis giving birth to live yound. Nothing is known concerning the number of endosymbionts during the growth cycle of the aphid. Using the quantitative polymerase chain reaction we will determine the number of copies of 16S and 23S rRNA genes in the B. aphidicola-aphid DNA preparations during the aphid growth cycle. The results will be an indication of the number of endosymbionts. Using immunological methods we will determine the amount of several essential endosymbiont consitutents. This includes ribosomes (ribosomal protein S1), RNA polymerase (beta-subunit), sigma factor and GroEL. %%% The results obtained from these studies may be of use for the eventual control of aphid populations by manipulation of the endosymbionts which are essential for their survival. ***