Department of Biological Sciences, Chungnam National University, Daejeon, Korea.
Division of Natural and Applied Sciences, Duke Kunshan University, Suzhou, China.
Plant Biol (Stuttg). 2023 Oct;25(6):915-923. doi: 10.1111/plb.13552. Epub 2023 Jul 1.
Reactive oxygen species (ROS) cause damage to various cellular processes in almost all organisms, in particular photosynthetic organisms that depend on the electron transfer chain for CO fixation. However, the detoxifying process to mitigate ROS damage has not been studied intensively in microalgae. Here, we characterized the ROS detoxifying role of a bZIP transcription factor, BLZ8, in Chlamydomonas reinhardtii. To identify downstream targets of BLZ8, we carried out comparative genome-wide transcriptomic profiling of BLZ8 OX and its parental CC-4533 under oxidative stress conditions. Luciferase reporter activity assays and RT-qPCR were performed to test whether BLZ8 regulates downstream genes. We performed an in silico functional gene network analysis and an in vivo immunoprecipitation assay to identify the interaction between downstream targets of BLZ8. Comparative transcriptomic analysis and RT-qPCR revealed that overexpression of BLZ8 increased the expression levels of plastid peroxiredoxin1 (PRX1) and ferredoxin-5 (FDX5) under oxidative stress conditions. BLZ8 alone could activate the transcriptional activity of FDX5 and required bZIP2 to activate transcriptional activity of PRX1. Functional gene network analysis using FDX5 and PRX1 orthologs in A. thaliana suggested that these two genes were functionally associated. Indeed, our immunoprecipitation assay revealed the physical interaction between PRX1 and FDX5. Furthermore, the complemented strain, fdx5 (FDX5), recovered growth retardation of the fdx5 mutant under oxidative stress conditions, indicating that FDX5 contributes to oxidative stress tolerance. These results suggest that BLZ8 activates PRX1 and FDX5 expression, resulting in the detoxification of ROS to confer oxidative stress tolerance in microalgae.
活性氧（ROS）会对几乎所有生物体的各种细胞过程造成损害，尤其是那些依赖电子传递链固定 CO 的光合生物。然而，在微藻中，ROS 解毒的过程并没有被深入研究。在这里，我们研究了莱茵衣藻 BLZ8 转录因子在 ROS 解毒中的作用。为了鉴定 BLZ8 的下游靶标，我们在氧化应激条件下对 BLZ8 OX 和其亲本 CC-4533 进行了全基因组比较转录组分析。我们进行了荧光素酶报告基因活性测定和 RT-qPCR 实验，以验证 BLZ8 是否调控下游基因。我们进行了计算机功能基因网络分析和体内免疫沉淀实验，以鉴定 BLZ8 的下游靶标之间的相互作用。比较转录组分析和 RT-qPCR 结果表明，BLZ8 的过表达在氧化应激条件下增加了质体过氧化物酶 1（PRX1）和铁氧还蛋白 5（FDX5）的表达水平。BLZ8 本身可以激活 FDX5 的转录活性，而需要 bZIP2 才能激活 PRX1 的转录活性。使用拟南芥的 FDX5 和 PRX1 同源基因进行功能基因网络分析表明，这两个基因在功能上是相关的。事实上，我们的免疫沉淀实验证实了 PRX1 和 FDX5 之间的物理相互作用。此外，补充了 FDX5 的突变株（FDX5）在氧化应激条件下恢复了 fdx5 突变株的生长迟缓，表明 FDX5 有助于提高对氧化应激的耐受性。这些结果表明，BLZ8 激活了 PRX1 和 FDX5 的表达，从而解毒 ROS，使微藻能够耐受氧化应激。
