小麦悬浮细胞应答激发子刺激的过敏性反应中Ca2+和 NO的动态变化及其相互作用

doi:10.3724/SP.J.1259.2015.00001

植物学报 ›› 2015, Vol. 50 ›› Issue (1): 1-11.DOI: 10.3724/SP.J.1259.2015.00001

• 研究报告 • 下一篇

小麦悬浮细胞应答激发子刺激的过敏性反应中Ca²⁺和 NO的动态变化及其相互作用

乔妹, 孙嘉炜, 陈琰, 韩胜芳, 侯春燕, 刘刚^*(), 王冬梅^*()

河北农业大学生命科学学院, 保定 071001

收稿日期:2013-10-30 接受日期:2014-02-23 出版日期:2015-01-01 发布日期:2015-04-09
通讯作者: 刘刚,王冬梅
作者简介:
? 共同第一作者
基金资助:
国家自然科学基金(No.31171472)、高等学校博士学科点专项科研基金(No.20111302130001)、河北省应用基础研究计划重点基础研究(No.08965505D)和河北省自然科学基金(No.C2007000515, No.C2010000787)

Dynamics and Interaction of Ca²⁺ and Nitric Oxide in Wheat Suspension Cells in the Hypersensitive Response

Mei Qiao, Jiawei Sun, Yan Chen, Shengfang Han, Chunyan Hou, Gang Liu^*, Dongmei Wang^*

College of Life Sciences, Agricultural University of Hebei, Baoding 071001, China

Received:2013-10-30 Accepted:2014-02-23 Online:2015-01-01 Published:2015-04-09
Contact: Liu Gang,Wang Dongmei
About author:
? These authors contributed equally to this paper

摘要/Abstract

摘要： 以感染叶锈菌的小麦(Triticum aestivum)叶片细胞间隙液IWF-260作为激发子, 刺激小麦品种洛夫林10和郑州5389的悬浮细胞, 探讨由激发子引发悬浮细胞过敏性反应中Ca²⁺和NO的变化及相互作用。以荧光分子探针Fluo-3AM和DAF-FM DA分别对细胞内Ca²⁺和NO进行标记, 利用激光共聚焦扫描显微镜对其动态变化进行实时监测, 通过药物学实验对Ca²⁺和NO的产生机制及其可能存在的相互关系进行探讨。结果表明, 2个小麦品种悬浮细胞的[Ca²⁺]_cyt水平对激发子刺激的反应表现出明显的差异, 对叶锈菌小种表现不亲和的洛夫林10悬浮细胞分别在激发子刺激后330秒和700秒出现2个钙峰; 而对该小种表现亲和的郑州5389悬浮细胞在激发子刺激后[Ca²⁺]_cyt水平稍有波动但变化不明显。药物学实验证明, [Ca²⁺]_cyt的升高依赖于胞外钙离子内流, 钙离子与激发子刺激诱发的过敏性防卫反应紧密相关。同样, 在激发子刺激后, 洛夫林10悬浮细胞出现1个NO峰, 而郑州5389悬浮细胞胞质NO变化不明显。药物学实验初步证明, NO的产生与胞外钙离子内流密切相关。由此推测, 在小麦悬浮细胞应答激发子刺激诱发的过敏性反应中, NO可能在钙的下游发挥作用。

Abstract: Suspension cells of wheat varieties Lovrin 10 and Zhengzhou 5389 were stimulated with IWF-260, a leaf intercellular washing fluid induced by leaf rust. The dynamics and interaction of Ca²⁺ and nitric oxide (NO) in the hypersensitive response induced by IWF-260 were analysed. The fluorescence molecular probe Fluo-3AM and DAF-FM DA were used to mark Ca²⁺ and NO, respectively. The two kinds of suspension cells showed differences in the concentration of Ca²⁺. Lovrin 10, a disease-resistant variety, showed two peaks of [Ca²⁺]_cyt, at 330 and 700 s, on stimulation, whereas Zhengzhou 5389, a susceptible variety, showed no obvious change in [Ca²⁺]_cyt with stimulation. The increased [Ca²⁺]_cyt depended on Ca²⁺ flowing into cells, which suggests that Ca²⁺ may be involved in the hypersensitive response. Like Ca²⁺, NO showed a similar pattern after elicitor stimulation. For Lovrin 10, NO showed a peak, with no change for Zhengzhou 5389. Thus, NO production and extracellular calcium influx are closely related to wheat suspension cells’ response to stimulation; NO may play a role downstream of calcium.

乔妹, 孙嘉炜, 陈琰, 韩胜芳, 侯春燕, 刘刚, 王冬梅. 小麦悬浮细胞应答激发子刺激的过敏性反应中Ca²⁺和 NO的动态变化及其相互作用. 植物学报, 2015, 50(1): 1-11.

Mei Qiao, Jiawei Sun, Yan Chen, Shengfang Han, Chunyan Hou, Gang Liu, Dongmei Wang. Dynamics and Interaction of Ca²⁺ and Nitric Oxide in Wheat Suspension Cells in the Hypersensitive Response. Chinese Bulletin of Botany, 2015, 50(1): 1-11.

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链接本文: https://www.chinbullbotany.com/CN/10.3724/SP.J.1259.2015.00001

https://www.chinbullbotany.com/CN/Y2015/V50/I1/1

图/表 4

图1 不同处理对小麦悬浮细胞死亡率的影响 (A) IWF对悬浮细胞死亡率的诱导; (B) LaCl3对IWF刺激诱发的悬浮细胞死亡率的影响; (C) EGTA对IWF刺激诱发的悬浮细胞死亡率的影响; (D) c-PTIO对IWF刺激诱发的悬浮细胞死亡率的影响

Figure 1 Effect of different treatments on the death rate of wheat suspension cell (A) The death rate of suspension cell induced by IWF; (B) Effect of LaCl3 on the death rate of suspension cell induced by IWF; (C) Effect of EGTA on the death rate of suspension cell induced by IWF; (D) Effect of c-PTIO on the death rate of suspension cell induced by IWF

图2 不同时间Fluo-3AM的装载效果 (A) 未装载Fluo-3AM的悬浮细胞; (B) 装载Fluo-3AM35分钟后的悬浮细胞; (C) 装载Fluo-3AM60分钟后的悬浮细胞。TR: 相应荧光图的透射光照片; 右下角色柱是本图所采用伪彩的色阶标尺。Bar=20 μm

Figure 2 The loading effect of Fluo-3AM under different time (A) Suspension cell of unloaded Fluo-3AM; (B) Suspension cell 35 mins after loading Fluo-3AM; (C) Suspension cell 60 mins after loading Fluo-3AM. TR: The transmission photo corresponding to the fluorescent. The color bar beside the image is the pseudocolor bar of this plate. Bar=20 μm

图3 小麦悬浮细胞经不同处理后[Ca2+]cyt的动态变化 (A) A23187和Mn2+处理后悬浮细胞[Ca2+]cyt的变化; (B) L10悬浮细胞经IWF处理后[Ca2+]cyt的动态变化; (C) 5389悬浮细胞经IWF处理后[Ca2+]cyt的动态变化; (D) EGTA和LaCl3处理对IWF260刺激L10悬浮细胞诱发的[Ca2+]cyt的影响; (E) c-PTIO处理对IWF260刺激L10悬浮细胞诱发的[Ca2+]cyt的影响

Figure 3 Cytosolic calcium dynamics in wheat suspension cell after different treatments (A) Changes of cytosolic calcium dynamics in suspension cell after A23187 and Mn2+ treatment; (B) Cytosolic calcium dynamics in L10 suspension cell after IWF treatment; (C) Cytosolic calcium dynamics in 5389 suspension cell after IWF treatment; (D) Effect of EGTA and LaCl3 on the cytosolic calcium dynamics in L10 suspension cell after IWF260 treatment; (E) Effect of c-PTIO on the cytosolic calcium dynamics in L10 suspension cell after IWF260 treatment

图4 小麦悬浮细胞经不同处理后胞质NO的动态变化 (A) L10悬浮细胞经SNP和c-PTIO处理后胞质NO动态变化; (B) L10悬浮细胞经IWF处理后胞质NO动态变化; (C) 5389悬浮细胞经IWF处理后胞质NO动态变化; (D) c-PTIO处理对IWF260刺激L10悬浮细胞诱发的胞质NO的影响; (E) EGTA和LaCl3处理对IWF260刺激L10悬浮细胞诱发的胞质NO的影响

Figure 4 Cytosolic NO dynamics in wheat suspension cell after different treatments (A) Cytosolic NO dynamics in L10 suspension cell after SNP and c-PTIO treatment; (B) Cytosolic NO dynamics in L10 suspension cell after IWF treatment; (C) Cytosolic NO dynamics in 5389 suspension cell after IWF treatment; (D) Effect of c-PTIO on cytosolic NO dynamics in L10 suspension cell after IWF260 treatment; (E) Effect of EGTA and LaCl3 on cytosolic NO dynamics in L10 suspension cell after IWF260 treatment

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小麦悬浮细胞应答激发子刺激的过敏性反应中Ca²⁺和 NO的动态变化及其相互作用

Dynamics and Interaction of Ca²⁺ and Nitric Oxide in Wheat Suspension Cells in the Hypersensitive Response

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