Kalanchoë PPC1 Is Essential for Crassulacean Acid Metabolism and the Regulation of Core Circadian Clock and Guard Cell Signaling Genes

ABSTRACTUnlike C3 plants, Crassulacean acid metabolism (CAM) plants fix CO2 in the dark using phosphoenolpyruvate carboxylase (PPC; EC 4.1.1.31). PPC combines PEP with CO2 (as HCO3−), forming oxaloacetate that is rapidly converted to malate, leading to vacuolar malic acid accumulation that peaks phased to dawn. In the light period, malate decarboxylation concentrates CO2 around RuBisCO for secondary fixation. CAM mutants lacking PPC have not been described. Here, RNAi was employed to silence 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 (LL) 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 were up-regulated and oscillated robustly. The regulation guard cell genes involved controlling stomatal movements was also altered in rPPC1-B. This provided direct evidence that altered regulatory patterns of key guard cell signaling genes are linked with the characteristic inverse pattern of stomatal opening and closing during CAM.

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Conservation and Divergence of Circadian Clock Operation in a Stress-Inducible Crassulacean Acid Metabolism Species Reveals Clock Compensation against Stress

PLANT PHYSIOLOGY ◽

10.1104/pp.104.054577 ◽

2005 ◽

Vol 137 (3) ◽

pp. 969-982 ◽

Cited By ~ 82

Author(s):

Susanna F. Boxall ◽

Jonathan M. Foster ◽

Hans J. Bohnert ◽

John C. Cushman ◽

Hugh G. Nimmo ◽

...

Keyword(s):

Circadian Clock ◽

Crassulacean Acid Metabolism ◽

Acid Metabolism

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Molecular dissection of the circadian clock output pathway that mediates the temporal coordination of Crassulacean acid metabolism

Comparative Biochemistry and Physiology Part A Molecular & Integrative Physiology ◽

10.1016/j.cbpa.2008.04.394 ◽

2008 ◽

Vol 150 (3) ◽

pp. S153

Author(s):

R. Meszter ◽

S. Boxall ◽

M. Jones ◽

J. Hartwell

Keyword(s):

Circadian Clock ◽

Crassulacean Acid Metabolism ◽

Acid Metabolism ◽

Molecular Dissection ◽

Temporal Coordination ◽

Circadian Clock Output

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Inference of Gene Regulatory Network Uncovers the Linkage Between Circadian Clock and Crassulacean Acid Metabolism in Kalanchoë fedtschenkoi

10.1101/745893 ◽

2019 ◽

Author(s):

Robert C. Moseley ◽

Francis Motta ◽

Gerald A. Tuskan ◽

Steve Haase ◽

Xiaohan Yang

Keyword(s):

Gene Expression ◽

Circadian Clock ◽

Gene Regulatory Network ◽

Regulatory Network ◽

Crassulacean Acid Metabolism ◽

Acid Metabolism ◽

Clock Genes ◽

Specific Gene ◽

Cam Plants ◽

Gene Regulatory

AbstractThe circadian clock drives time-specific gene expression, allowing for associated biological processes to be active during certain times of the 24 h day. Crassulacean acid metabolism (CAM) photosynthetic plants represent an interesting case of circadian regulation of gene expression as CO2 fixation and stomatal movement in CAM plants display strong circadian dynamics. The molecular mechanisms behind how the circadian clock enabled these physiological differences is not well understood. Therefore, we set out to investigate whether core circadian elements in CAM plants were re-phased during evolution, or whether networks of phase-specific genes were simply connected to different core elements. We utilized a new metric for identifying candidate core genes of a periodic gene network and then applied the Local Edge Machine (LEM) algorithm to infer regulatory relationships between the candidate core clock genes and orthologs of known core clock genes in K. fedtschenkoi. We also used LEM to identify stomata-related gene targets for K. fedtschenkoi core clock genes and constructed a subsequent gene regulatory network. Our results provide new insights into the mechanism of circadian control of CAM-related genes in K. fedtschenkoi, facilitating the engineering of CAM machinery into non-CAM plants for sustainable crop production in water-limited environments.

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Inference of Gene Regulatory Network Uncovers the Linkage between Circadian Clock and Crassulacean Acid Metabolism in Kalanchoë fedtschenkoi

Cells ◽

10.3390/cells10092217 ◽

2021 ◽

Vol 10 (9) ◽

pp. 2217

Author(s):

Robert C. Moseley ◽

Francis Motta ◽

Gerald A. Tuskan ◽

Steven B. Haase ◽

Xiaohan Yang

Keyword(s):

Gene Expression ◽

Circadian Clock ◽

Gene Regulatory Network ◽

Regulatory Network ◽

Crassulacean Acid Metabolism ◽

Acid Metabolism ◽

Clock Genes ◽

Stomatal Movement ◽

Cam Plants ◽

Gene Regulatory

The circadian clock drives time-specific gene expression, enabling biological processes to be temporally controlled. Plants that conduct crassulacean acid metabolism (CAM) photosynthesis represent an interesting case of circadian regulation of gene expression as stomatal movement is temporally inverted relative to stomatal movement in C3 plants. The mechanisms behind how the circadian clock enabled physiological differences at the molecular level is not well understood. Recently, the rescheduling of gene expression was reported as a mechanism to explain how CAM evolved from C3. Therefore, we investigated whether core circadian clock genes in CAM plants were re-phased during evolution, or whether networks of phase-specific genes were simply re-wired to different core clock genes. We identified candidate core clock genes based on gene expression features and then applied the Local Edge Machine (LEM) algorithm to infer regulatory relationships between this new set of core candidates and known core clock genes in Kalanchoë fedtschenkoi. We further inferred stomata-related gene targets for known and candidate core clock genes and constructed a gene regulatory network for core clock and stomata-related genes. Our results provide new insight into the mechanism of circadian control of CAM-related genes in K. fedtschenkoi, facilitating the engineering of CAM machinery into non-CAM plants for sustainable crop production in water-limited environments.

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