Abscisic acid (ABA)-activated inward Ca2+-permeable channels in the plasma membrane (PM) of guard cells are required for the initiation and regulation of ABA-specific cytosolic Ca2+ signaling and stomatal closure in plants. But the identities of the PM Ca2+ channels are still unknown. We hypothesized that the ABA-activated Ca2+ channels consist of multiple CYCLIC NUCLEOTIDE-GATED CHANNEL (CNGC) proteins from the CNGC family, which is known as a Ca2+-permeable channel family in Arabidopsis (Arabidopsis thaliana). In this research, we observed high expression of multiple CNGC genes in Arabidopsis guard cells, namely CNGC5, CNGC6, CNGC9, and CNGC12. The T-DNA insertional loss-of-function quadruple mutant cngc5-1 cngc6-2 cngc9-1 cngc12-1 (hereafter c5/6/9/12) showed a strong ABA-insensitive phenotype of stomatal closure. Further analysis revealed that ABA-activated Ca2+ channel currents were impaired, and ABA-specific cytosolic Ca2+ oscillation patterns were disrupted in c5/6/9/12 guard cells compared with in wild-type guard cells. All ABA-related phenotypes of the c5/6/9/12 mutant were successfully rescued by the expression of a single gene out of the four CNGCs under the respective native promoter. Thus, our findings reveal a type of ABA-activated PM Ca2+ channel comprising multiple CNGCs, which is essential for ABA-specific Ca2+ signaling of guard cells and ABA-induced stomatal closure in Arabidopsis.

中文翻译：

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拟南芥中多个环核苷酸门控通道充当 ABA 激活的 Ca2+ 通道，是 ABA 诱导的气孔关闭所需的。

保卫细胞质膜 (PM) 中脱落酸 (ABA) 激活的内向 Ca2+ 通透通道是植物中 ABA 特异性胞质 Ca2+ 信号传导和气孔关闭的启动和调节所必需的。但 PM Ca2+ 通道的身份仍然未知。我们假设 ABA 激活的 Ca2+ 通道由来自 CNGC 家族的多个环状核苷酸门控通道 (CNGC) 蛋白组成，该家族在拟南芥 (Arabidopsis thaliana) 中被称为 Ca2+ 通透性通道家族。在本研究中，我们观察到拟南芥保卫细胞中多个CNGC基因的高表达，即CNGC5、CNGC6、CNGC9和CNGC12。T-DNA插入功能丧失四重突变体cngc5-1 cngc6-2 cngc9-1 cngc12-1（以下简称c5/6/9/12）表现出强烈的ABA不敏感的气孔关闭表型。进一步分析表明，与野生型保卫细胞相比，c5/6/9/12 保卫细胞中 ABA 激活的 Ca2+ 通道电流受损，ABA 特异性胞质 Ca2+ 振荡模式被破坏。c5/6/9/12 突变体的所有 ABA 相关表型均通过在各自的天然启动子下表达四个 CNGC 中的单个基因而成功挽救。因此，我们的研究结果揭示了一种由 ABA 激活的 PM Ca2+ 通道，包含多个 CNGC，这对于拟南芥中保卫细胞的 ABA 特异性 Ca2+ 信号传导和 ABA 诱导的气孔关闭至关重要。