Liu Kaiwei, Zhao Huan, Lee Keun Pyo, Yu Qing, Di Minghui, Wang Liangsheng, Kim Chanhong
Shanghai Center for Plant Stress Biology, CAS Center for Excellence in Molecular Plant Sciences (CEMPS), Chinese Academy of Sciences, Shanghai 200032, China.
State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing 100193, China.
Plant Cell. 2024 Dec 23;37(1). doi: 10.1093/plcell/koae296.
Chloroplasts are recognized as environmental sensors, capable of translating environmental fluctuations into diverse signals to communicate with the nucleus. Among the reactive oxygen species produced in chloroplasts, singlet oxygen (1O2) has been extensively studied due to its dual roles, encompassing both damage and signaling activities, and the availability of conditional mutants overproducing 1O2 in chloroplasts. In particular, investigating the Arabidopsis (Arabidopsis thaliana) mutant known as fluorescent (flu) has led to the discovery of EXECUTER1 (EX1), a plastid 1O2 sensor residing in the grana margin of the thylakoid membrane. 1O2-triggered EX1 degradation is critical for the induction of 1O2-responsive nuclear genes (SOrNGs). However, a recent study showed that EX1 relocates from chloroplasts to the nucleus upon 1O2 release, where it interacts with WRKY18 and WRKY40 (WRKY18/40) transcription factors to regulate SOrNG expression. In this study, we challenge this assertion. Our confocal microscopy analysis and subcellular fractionation assays demonstrate that EX1 does not accumulate in the nucleus. While EX1 appears in nuclear fractions, subsequent thermolysin treatment assays indicate that it adheres to the outer nuclear region rather than localizing inside the nucleus. Furthermore, luciferase complementation imaging and yeast 2-hybrid assays reveal that EX1 does not interact with nuclear WRKY18/40. Consequently, our study refines the current model of 1O2 signaling by ruling out the nuclear relocation of intact EX1 as a means of communication between the chloroplast and nucleus.
叶绿体被认为是环境传感器，能够将环境波动转化为多种信号与细胞核进行通信。在叶绿体产生的活性氧中，单线态氧（1O2）因其具有损伤和信号传导双重作用以及叶绿体中过量产生1O2的条件性突变体的可得性而得到广泛研究。特别是，对拟南芥（Arabidopsis thaliana）中名为fluorescent（flu）的突变体的研究导致发现了EXECUTER1（EX1），一种位于类囊体膜基粒边缘的质体1O2传感器。1O2触发的EX1降解对于诱导1O2响应性核基因（SOrNGs）至关重要。然而，最近的一项研究表明，1O2释放后EX1从叶绿体重新定位到细胞核，在那里它与WRKY18和WRKY40（WRKY18/40）转录因子相互作用以调节SOrNG表达。在本研究中，我们对这一观点提出质疑。我们的共聚焦显微镜分析和亚细胞分级分离试验表明EX1不会在细胞核中积累。虽然EX1出现在细胞核分级分离物中，但随后的嗜热菌蛋白酶处理试验表明它附着在细胞核外部区域而非定位于细胞核内部。此外，荧光素酶互补成像和酵母双杂交试验表明EX1不会与细胞核中的WRKY18/40相互作用。因此，我们的研究通过排除完整的EX1核重新定位作为叶绿体与细胞核之间通信方式，完善了当前的1O2信号传导模型。
