College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China.
College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, People's Republic of China; Jiangsu Collaborative Innovation Center for Solid Organic Waste Resource, Nanjing Agricultural University, Nanjing 210095, People's Republic of China.
Aquat Toxicol. 2020 Sep;226:105585. doi: 10.1016/j.aquatox.2020.105585. Epub 2020 Jul 28.
Although excess ammoniacal-nitrogen (NH-N) results in the disturbance of various important biochemical and physiological processes, a detailed study on the effects of NH-N stress on the photosynthesis and global changes in protein levels in submerged macrophytes is still lacking. Here, the changes of excess NH-N on physiological parameters in Hydrilla verticillata (L.f.) Royle, a submerged macrophyte were investigated, including the contents of photosynthetic pigments, soluble sugars, net photosynthesis and respiration, glutamine synthetase (GS) and glutamate synthase (GOGAT) activities, chloroplast ultrastructure, chloroplast reactive oxygen species (ROS) accumulation and protein levels. Our results showed that the net photosynthetic rate and pigment content reached maximum values when the plants were treated with 1 and 2 mg L NH-N, respectively, and decreased at NH-N concentrations at 5, 10, 15 and 20 mg L. This decrease might be caused by ROS accumulation. Compared that in 0.02 mg L NH-N as a control, ROS generation in chloroplasts significantly increased in the presence of more than 2 mg L NH-N. Consistently, the damages caused by over-accumulated ROS were observed in chloroplast ultrastructure, showing a loose thylakoid membranes and swollen grana/stroma lamellae. Furthermore, through proteomic analysis, we identified 91 differentially expressed protein spots. Among them, six proteins involved in photosynthesis decreased in abundance in response to excess NH-N. Surprisingly, the abundance of all the identified proteins that were involved in nitrogen assimilation and amino acid metabolism tended to increase under excess NH-N compared with the control, suggestive of the imbalanced carbon and nitrogen (C-N) metabolisms. In support, activated GS and GOGAT cycle was observed, evidenced by higher activities of GS and GOGAT enzymes. To our knowledge, this work is the first description that excess NH-N results in chloroplast ultrastructural damages and the first proteomic evidence to support that excess NH-N can lead to a decline in photosynthesis and imbalance of C-N metabolism in submerged macrophytes.
虽然过量的氨氮（NH-N）会导致各种重要的生化和生理过程紊乱，但对于 NH-N 胁迫对沉水植物光合作用和整体蛋白质水平变化的影响，仍缺乏详细的研究。在这里，研究了过量 NH-N 对沉水植物水鳖（L.f.）Royale 的生理参数的影响，包括光合色素、可溶性糖、净光合作用和呼吸作用、谷氨酰胺合成酶（GS）和谷氨酸合酶（GOGAT）活性、叶绿体超微结构、叶绿体活性氧（ROS）积累和蛋白质水平的变化。我们的结果表明，当植物分别用 1 和 2 mg L NH-N 处理时，净光合速率和色素含量达到最大值，而在 NH-N 浓度为 5、10、15 和 20 mg L 时则降低。这种降低可能是由于 ROS 积累造成的。与 0.02 mg L NH-N 作为对照相比，当 NH-N 浓度超过 2 mg L 时，叶绿体中 ROS 的产生显著增加。一致地，在叶绿体超微结构中观察到由于 ROS 过度积累而引起的损伤，表现为疏松的类囊体膜和肿胀的基粒/基质片层。此外，通过蛋白质组学分析，我们鉴定了 91 个差异表达的蛋白点。其中，有 6 个参与光合作用的蛋白丰度因过量 NH-N 而降低。令人惊讶的是，与对照相比，所有参与氮同化和氨基酸代谢的鉴定蛋白的丰度在过量 NH-N 下都趋于增加，表明碳氮（C-N）代谢的不平衡。支持这一观点的是，观察到激活的 GS 和 GOGAT 循环，GS 和 GOGAT 酶的活性更高。据我们所知，这是首次描述过量 NH-N 导致叶绿体超微结构损伤的工作，也是首次通过蛋白质组学证据支持过量 NH-N 会导致沉水植物光合作用下降和 C-N 代谢失衡的证据。
