Table_5_The Puzzle of Metabolite Exchange and Identification of Putative Octotrico Peptide Repeat Expression Regulators in the Nascent Photosynthetic Organelles of Paulinella chromatophora.XLSX

The endosymbiotic acquisition of mitochondria and plastids more than one billion years ago was central for the evolution of eukaryotic life. However, owing to their ancient origin, these organelles provide only limited insights into the initial stages of organellogenesis. The cercozoan amoeba Paulinella chromatophora contains photosynthetic organelles—termed chromatophores—that evolved from a cyanobacterium ∼100 million years ago, independently from plastids in plants and algae. Despite the more recent origin of the chromatophore, it shows tight integration into the host cell. It imports hundreds of nucleus-encoded proteins, and diverse metabolites are continuously exchanged across the two chromatophore envelope membranes. However, the limited set of chromatophore-encoded solute transporters appears insufficient for supporting metabolic connectivity or protein import. Furthermore, chromatophore-localized biosynthetic pathways as well as multiprotein complexes include proteins of dual genetic origin, suggesting that mechanisms evolved that coordinate gene expression levels between chromatophore and nucleus. These findings imply that similar to the situation in mitochondria and plastids, also in P. chromatophora nuclear factors evolved that control metabolite exchange and gene expression in the chromatophore. Here we show by mass spectrometric analyses of enriched insoluble protein fractions that, unexpectedly, nucleus-encoded transporters are not inserted into the chromatophore inner envelope membrane. Thus, despite the apparent maintenance of its barrier function, canonical metabolite transporters are missing in this membrane. Instead we identified several expanded groups of short chromatophore-targeted orphan proteins. Members of one of these groups are characterized by a single transmembrane helix, and others contain amphipathic helices. We hypothesize that these proteins are involved in modulating membrane permeability. Thus, the mechanism generating metabolic connectivity of the chromatophore fundamentally differs from the one for mitochondria and plastids, but likely rather resembles the poorly understood mechanism in various bacterial endosymbionts in plants and insects. Furthermore, our mass spectrometric analysis revealed an expanded family of chromatophore-targeted helical repeat proteins. These proteins show similar domain architectures as known organelle-targeted expression regulators of the octotrico peptide repeat type in algae and plants. Apparently these chromatophore-targeted proteins evolved convergently to plastid-targeted expression regulators and are likely involved in gene expression control in the chromatophore.

超过十亿年前，线粒体和质体的共生获取对真核生物的进化具有核心意义。然而，由于这些细胞器的古老起源，它们只能提供有限的见解来探索细胞器起源的初期阶段。围网虫类变形虫保罗宁拉·色谱体（Paulinella chromatophora）含有光合细胞器，被称为色素体，这些色素体大约在一亿年前从蓝细菌进化而来，独立于植物和藻类中的质体。尽管色素体的起源较晚，但它与宿主细胞的整合却非常紧密。它导入数百个由细胞核编码的蛋白质，并且多种代谢物在两个色素体包膜膜之间持续交换。然而，色素体编码的溶质转运蛋白的有限集合似乎不足以支持代谢连通性或蛋白质的导入。此外，色素体定位的合成途径以及多蛋白复合体包括具有双遗传起源的蛋白质，这表明已演化出协调色素体与细胞核之间基因表达水平的机制。这些发现表明，与线粒体和质体的情形类似，在P. 色谱体中，细胞核因子也演化出来以控制色素体中的代谢物交换和基因表达。在本研究中，我们通过富含不溶性蛋白分数的质谱分析发现，出人意料的是，细胞核编码的转运蛋白并未插入色素体内部包膜膜。因此，尽管看似维持了其屏障功能，但经典代谢物转运蛋白却缺失于该膜中。相反，我们识别了几个扩展的短色素体靶向孤儿蛋白群体。其中一组蛋白的特征是具有单个跨膜螺旋，而其他则包含两亲性螺旋。我们假设这些蛋白参与调节膜通透性。因此，生成色素体代谢连通性的机制与线粒体和质体截然不同，而更类似于植物和昆虫中各种细菌共生体中尚不为人知的机制。此外，我们的质谱分析揭示了扩展的色素体靶向螺旋重复蛋白家族。这些蛋白显示出与已知的海藻和植物中八肽重复型细胞器靶向表达调控因子相似的域结构。显然，这些色素体靶向蛋白在进化上趋同于质体靶向表达调控因子，并可能在色素体的基因表达调控中发挥作用。