Cadmium-Induced Hydrogen Sulfide Synthesis Is Involved in Cadmium Tolerance in Medicago sativa by Reestablishment of Reduced (Homo)glutathione and Reactive Oxygen Species Homeostases

Until now, physiological mechanisms and downstream targets responsible for the cadmium (Cd) tolerance mediated by endogenous hydrogen sulfide (H2S) have been elusive. To address this gap, a combination of pharmacological, histochemical, biochemical and molecular approaches was applied. The perturbation of reduced (homo)glutathione homeostasis and increased H2S production as well as the activation of two H2S-synthetic enzymes activities, including L-cysteine desulfhydrase (LCD) and D-cysteine desulfhydrase (DCD), in alfalfa seedling roots were early responses to the exposure of Cd. The application of H2S donor sodium hydrosulfide (NaHS), not only mimicked intracellular H2S production triggered by Cd, but also alleviated Cd toxicity in a H2S-dependent fashion. By contrast, the inhibition of H2S production caused by the application of its synthetic inhibitor blocked NaHS-induced Cd tolerance, and destroyed reduced (homo)glutathione and reactive oxygen species (ROS) homeostases. Above mentioned inhibitory responses were further rescued by exogenously applied glutathione (GSH). Meanwhile, NaHS responses were sensitive to a (homo)glutathione synthetic inhibitor, but reversed by the cotreatment with GSH. The possible involvement of cyclic AMP (cAMP) signaling in NaHS responses was also suggested. In summary, LCD/DCD-mediated H2S might be an important signaling molecule in the enhancement of Cd toxicity in alfalfa seedlings mainly by governing reduced (homo)glutathione and ROS homeostases.

迄今为止，由内源性硫化氢（endogenous hydrogen sulfide, H2S）介导的镉（cadmium, Cd）耐受相关的生理机制与下游靶点仍不明确。为填补这一研究空白，本研究联合运用了药理学、组织化学、生物化学与分子生物学实验手段。研究发现，苜蓿幼苗根系中还原型（同型）谷胱甘肽（reduced (homo)glutathione）稳态失衡、硫化氢生成增加，以及两种硫化氢合成酶——L-半胱氨酸脱巯基酶（L-cysteine desulfhydrase, LCD）与D-半胱氨酸脱巯基酶（D-cysteine desulfhydrase, DCD）——的活性被激活，上述均为植株暴露于镉胁迫后的早期响应事件。施加硫化氢供体硫氢化钠（sodium hydrosulfide, NaHS），不仅可模拟镉诱导的胞内硫化氢生成，还能以硫化氢依赖的方式缓解镉毒性。与之相反，使用硫化氢合成抑制剂阻断硫化氢生成后，不仅会抵消硫氢化钠诱导的镉耐受效应，还会破坏还原型（同型）谷胱甘肽与活性氧（reactive oxygen species, ROS）的稳态平衡。外源性施加谷胱甘肽（glutathione, GSH）可进一步挽救上述抑制性表型。同时，硫氢化钠介导的调控过程对（同型）谷胱甘肽合成抑制剂敏感，而与谷胱甘肽共处理则可逆转这一敏感性。研究还提示，环磷酸腺苷（cyclic AMP, cAMP）信号通路可能参与了硫氢化钠的调控过程。综上，LCD/DCD介导的硫化氢可能是苜蓿幼苗缓解镉毒性过程中的重要信号分子，其核心作用机制为调控还原型（同型）谷胱甘肽与活性氧的稳态平衡。