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Phosphatidic acid (PtdOH) is emerging as an important signaling lipid in abiotic stress responses in plants. The effect of cold stress was monitored using  32 P-labeled seedlings and leaf discs of  Arabidopsis thaliana . Low, non-freezing temperatures were found to trigger a very rapid  32 P-PtdOH increase, peaking within 2 and 5 min, respectively. In principle, PtdOH can be generated through three different pathways, i.e., (1) via  de novo  phospholipid biosynthesis (through acylation of lyso-PtdOH), (2) via phospholipase D hydrolysis of structural phospholipids, or (3) via phosphorylation of diacylglycerol (DAG) by DAG kinase (DGK). Using a differential  32 P-labeling protocol and a PLD-transphosphatidylation assay, evidence is provided that the rapid  32 P-PtdOH response was primarily generated through DGK. A simultaneous decrease in the levels of  32 P-PtdInsP, correlating in time, temperature dependency, and magnitude with the increase in  32 P-PtdOH, suggested that a PtdInsP-hydrolyzing PLC generated the DAG in this reaction. Testing T-DNA insertion lines available for the seven  DGK  genes, revealed no clear changes in  32 P-PtdOH responses, suggesting functional redundancy. Similarly, known cold-stress mutants were analyzed to investigate whether the PtdOH response acted downstream of the respective gene products. The  hos1, los1 , and  fry1  mutants were found to exhibit normal PtdOH responses. Slight changes were found for  ice1, snow1 , and the overexpression line  Super-ICE1 , however, this was not cold-specific and likely due to pleiotropic effects. A tentative model illustrating direct cold effects on phospholipid metabolism is presented.

Rapid phosphatidic acid accumulation in response to low temperature stress in Arabidopsis is generated through diacylglycerol kinase