Lamotte Olivier, Courtois Cécile, Dobrowolska Grazyna, Besson Angélique, Pugin Alain, Wendehenne David
Unité Mixte de Recherche INRA 1088/CNRS 5184/Université de Bourgogne, Plante-Microbe-Environnement, Dijon, France.
Free Radic Biol Med. 2006 Apr 15;40(8):1369-76. doi: 10.1016/j.freeradbiomed.2005.12.006. Epub 2006 Jan 6.
In this study, we investigated a role for nitric oxide (NO) in mediating the elevation of the free cytosolic Ca(2+) concentration (Ca(2+)) in plants using Nicotiana plumbaginifolia cells expressing the Ca(2+) reporter apoaequorin. Hyperosmotic stress induced a fast increase of Ca(2+) which was strongly reduced by pretreating cell suspensions with the NO scavenger carboxy PTIO, indicating that NO mediates Ca(2+) changes in plant cells challenged by abiotic stress. Accordingly, treatment of transgenic N. plumbaginifolia cells with the NO donor diethylamine NONOate was followed by a transient increase of Ca(2+) sensitive to plasma membrane Ca(2+) channel inhibitors and antagonist of cyclic ADP ribose. We provided evidence that NO might activate plasma membrane Ca(2+) channels by inducing a rapid and transient plasma membrane depolarization. Furthermore, NO-induced elevation of Ca(2+) was suppressed by the kinase inhibitor staurosporine, suggesting that NO enhances Ca(2+) by promoting phosphorylation-dependent events. This result was further supported by the demonstration that the NO donor induced the activation of a 42-kDa protein kinase which belongs to SnRK2 families and corresponds to Nicotiana tabacum osmotic-stress-activated protein kinase (NtOSAK). Interestingly, NtOSAK was activated in response to hyperosmotic stress through a NO-dependent process, supporting the hypothesis that NO also promotes protein kinase activation during physiological processes.
在本研究中，我们利用表达钙离子报告蛋白脱辅基水母发光蛋白的烟草细胞，研究了一氧化氮（NO）在介导植物细胞溶质游离钙离子浓度（[Ca²⁺]cyt）升高过程中的作用。高渗胁迫诱导[Ca²⁺]cyt快速升高，而用NO清除剂羧基-PTIO预处理细胞悬液可显著降低这种升高，这表明NO介导了非生物胁迫刺激下植物细胞中[Ca²⁺]cyt的变化。相应地，用NO供体二乙胺NONOate处理转基因烟草细胞后，[Ca²⁺]cyt会短暂升高，且对质膜钙离子通道抑制剂和环ADP核糖拮抗剂敏感。我们提供的证据表明，NO可能通过诱导质膜快速且短暂的去极化来激活质膜钙离子通道。此外，激酶抑制剂星形孢菌素可抑制NO诱导的[Ca²⁺]cyt升高，这表明NO通过促进磷酸化依赖事件来增强[Ca²⁺]cyt。NO供体诱导一种属于SnRK2家族且与烟草渗透胁迫激活蛋白激酶（NtOSAK）相对应的42 kDa蛋白激酶活化，进一步支持了这一结果。有趣的是，NtOSAK在高渗胁迫下通过NO依赖过程被激活，支持了NO在生理过程中也促进蛋白激酶活化的假说。
