Eggenberger Kai, Sanyal Papia, Hundt Svenja, Wadhwani Parvesh, Ulrich Anne S, Nick Peter
Botanical Institute and DFG-Center of Functional Nanostructures (CFN), Karlsruhe Institute of Technology (KIT), Kaiserstr, Karlsruhe, Germany.
Institute of Biological Interfaces (IBG-2) and DFG-Center of Functional Nanostructures (CFN), Institute of Organic Chemistry, Karlsruhe Institute of Technology (KIT), Fritz-Haber Weg 6, Karlsruhe, Germany.
Plant Cell Physiol. 2017 Jan 1;58(1):71-85. doi: 10.1093/pcp/pcw161.
Actin filaments are essential for the integrity of the cell membrane. In addition to this structural role, actin can modulate signaling by altering polar auxin flow. On the other hand, the organization of actin filaments is modulated by auxin constituting a self-referring signaling hub. Although the function of this auxin–actin oscillator is not clear, there is evidence for a functional link with stress signaling activated by the NADPH oxidase Respiratory burst oxidase Homolog (RboH). In the current work, we used the cell-penetrating peptide BP100 to induce a mild and transient perturbation of membrane integrity. We followed the response of actin to the BP100 uptake in a green fluorescent protein (GFP)-tagged actin marker line of tobacco Bright Yellow 2 (BY-2) cells by spinning disc confocal microscopy. We observed that BP100 enters in a stepwise manner and reduces the extent of actin remodeling. This actin ‘freezing’ can be rescued by the natural auxin IAA, and mimicked by the auxin-efflux inhibitor 1-napthylphthalamic acid (NPA). We further tested the role of the membrane-localized NADPH oxidase RboH using the specific inhibitor diphenyl iodonium (DPI), and found that DPI acts antagonistically to BP100, although DPI alone can induce a similar actin ‘freezing’ as well. We propose a working model, where the mild violation of membrane integrity by BP100 stimulates RboH, and the resulting elevated levels of reactive oxygen species interfere with actin dynamicity. The mitigating effect of auxin is explained by competition of auxin- and RboH-triggered signaling for superoxide anions. This self-referring auxin–actin–RboH hub might be essential for integrity sensing.
肌动蛋白丝对于细胞膜的完整性至关重要。除了这一结构作用外，肌动蛋白还可通过改变极性生长素流来调节信号传导。另一方面，肌动蛋白丝的组织受到生长素的调节，构成了一个自我参照的信号枢纽。尽管这种生长素 - 肌动蛋白振荡器的功能尚不清楚，但有证据表明它与由NADPH氧化酶呼吸爆发氧化酶同源物（RboH）激活的应激信号存在功能联系。在当前的工作中，我们使用细胞穿透肽BP100诱导细胞膜完整性的轻度和短暂扰动。我们通过转盘共聚焦显微镜观察了烟草Bright Yellow 2（BY-2）细胞中绿色荧光蛋白（GFP）标记的肌动蛋白标记系中肌动蛋白对BP100摄取的反应。我们观察到BP100以逐步方式进入并减少肌动蛋白重塑的程度。这种肌动蛋白“冻结”可以被天然生长素IAA挽救，并被生长素外流抑制剂1-萘基邻苯二甲酸（NPA）模拟。我们进一步使用特异性抑制剂二苯基碘鎓（DPI）测试了膜定位的NADPH氧化酶RboH的作用，发现DPI与BP100起拮抗作用，尽管单独的DPI也可以诱导类似的肌动蛋白“冻结”。我们提出了一个工作模型，其中BP100对膜完整性的轻度破坏刺激了RboH，并且由此产生的活性氧水平升高干扰了肌动蛋白的动态性。生长素的缓解作用通过生长素和RboH触发的信号对超氧阴离子的竞争来解释。这种自我参照的生长素 - 肌动蛋白 - RboH枢纽可能对完整性感知至关重要。
