Unlike C₃ plants, Crassulacean acid metabolism (CAM) plants fix CO₂ in the dark using phosphoenolpyruvate carboxylase (PPC; EC 4.1.1.31). PPC combines phosphoenolpyruvate with CO₂ (as HCO₃^–), forming oxaloacetate. The oxaloacetate is converted to malate, leading to malic acid accumulation in the vacuole, which peaks at dawn. During the light period, malate decarboxylation concentrates CO₂ around Rubisco for secondary fixation. CAM mutants lacking PPC have not been described. Here, we employed RNA interference to silence the CAM isogene PPC1 in Kalanchoë laxiflora. Line rPPC1-B lacked PPC1 transcripts, PPC activity, dark period CO₂ fixation, and nocturnal malate accumulation. Light period stomatal closure was also perturbed, and the plants displayed reduced but detectable dark period stomatal conductance and arrhythmia of the CAM CO₂ fixation circadian rhythm under constant light and temperature free-running conditions. By contrast, the rhythm of delayed fluorescence was enhanced in plants lacking PPC1. Furthermore, a subset of gene transcripts within the central circadian oscillator was upregulated and oscillated robustly in this line. The regulation of guard cell genes involved in controlling stomatal movements was also perturbed in rPPC1-B. These findings provide direct evidence that the regulatory patterns of key guard cell signaling genes are linked with the characteristic inverse pattern of stomatal opening and closing during CAM.

与C ₃植物不同，Crassulacean酸代谢（CAM）植物在黑暗中使用磷酸烯醇丙酮酸羧化酶（PPC; EC 4.1.1.31）固定CO ₂。PPC将磷酸烯醇丙酮酸与CO ₂（作为HCO ₃ ^–）结合，形成草酰乙酸。草酰乙酸被转化为苹果酸，导致苹果酸在液泡中积累，并在黎明时达到峰值。在光照期间，苹果酸脱羧使Rubisco周围的CO ₂集中，以进行二次固定。缺乏PPC的CAM突变体尚未描述。在这里，我们采用RNA干扰技术沉默CAM同源基因PPC1在长寿花水柏枝。线rPPC1-B缺少PPC1转录本，PPC活性，暗期CO ₂固定和夜间苹果酸积累。光照期气孔关闭也受到干扰，并且在恒定的光照和温度自由运行条件下，植物显示出减少但可检测到的黑暗期气孔导度和CAM CO ₂固定昼夜节律的心律不齐。相比之下，在缺乏PPC1的植物中，延迟荧光的节奏增强了。此外，在中央昼夜节律振荡器中的基因转录子的一个子集在这一行中被上调并强烈振荡。rPPC1-B也干扰了参与控制气孔运动的保卫细胞基因的调控。这些发现提供了直接的证据，表明关键的保卫细胞信号转导基因的调控模式与CAM期间气孔打开和关闭的特征逆模式有关。