Mechanisms and regulation of intracellular iron trafficking in Chlamydomonas reinhardtii

铁是所有光合生物的重要微量营养素。莱茵衣藻（Chlamydiumreinhardtii）通过高亲和力摄取系统同化铁，所述高亲和力摄取系统包括铁还原酶（FRE 1）和铁氧化酶-铁通透酶系统（FOX 1-FTR 1）。此外，C. reinhardtii细胞通过IRT 1/IRT 2转运系统同化铁（和可能的其它金属）。虽然已经研究并建立了铁输入到藻类细胞中的机制，但铁如何在细胞质和细胞器之间运输以及如何调节其同化和运输的机制目前尚不清楚。缺铁C. reinhardtii鉴定了用于细胞内铁运输和调节的几种候选物的转录水平增加。我们推测，这些蛋白质在细胞器之间的铁的运输和铁稳态的调节中发挥重要作用。通过反向遗传学和生物化学方法相结合，我们将分析这些蛋白质在C。莱因哈德氏菌同时，将采用转录组学（RNA-Seq）、蛋白质组学（LC-MS/MS）和金属含量测定（ICP-MS）对来自这些蛋白质的野生型和敲低突变体的全细胞和分离细胞器进行分析。这将使我们更好地了解细胞内铁的分布和铁稳态的调节，在C。莱因哈德氏菌此外，我们还将采用pull-down分析来鉴定调控C.莱因哈德氏菌

Iron is an important micronutrient for all photosynthetic organisms. Chlamydomonas reinhardtii, assimilate iron via a high affinity uptake system which comprises a ferric-reductase (FRE1) and a ferroxidase-ferric-permease system (FOX1-FTR1). Additionally, C. reinhardtii cells assimilate iron (and possibly other metals) via IRT1/IRT2 transporter system. While mechanisms of iron import into algal cells have been studied and established, mechanisms of how iron is transported between cytoplasm and organelles and how its assimilation and trafficking are regulated are currently unknown. Recent transcriptome analysis of iron-deficient C. reinhardtii identified increased transcript levels of several candidates for intracellular iron trafficking and regulation. We hypothesize that these proteins play an important role in the transport of iron between organelles and in the regulation of iron homeostasis. By combining reverse genetics and biochemical approaches we will analyze the function of these proteins in C. reinhardtii. In parallel, transcriptomics (RNA-Seq), proteomics (LC-MS/MS) and determination of metal content (ICP-MS) on whole cells and isolated organelles from wild-type and knock-down mutants of these proteins will be employed. This will allow us to better understand the intracellular iron distribution and the regulation of iron homeostasis in C. reinhardtii. In addition, a pull-down assay will be employed with the aim to identify the transcription factor responsible for the regulation of iron deficiency responses in C. reinhardtii.