Transcriptional and post-translational upregulation of phosphoenolpyruvate carboxylase in Arabidopsis thaliana (L. Heynh) under cadmium stress

Phosphoenolpyruvate carboxylase (PEPC) is a tightly controlled anaplerotic enzyme that replenishes organic acids that have been withdrawn from the tricarboxylic acid cycle. Organic acid anions are important chelators of toxic metals, including cadmium (Cd). To determine the Cd-stress responses of two isozymes of PEPC in Arabidopsis (AtPPC1 and AtPPC3), we compared wild type (Col-0) with two T-DNA insertional mutant lines (atppc1 and atppc3). Plant mass decreased in response to Cd in all lines, but atppc3 was the most sensitive to Cd even though its roots contained 30% lower concentrations of Cd. Treatment with 1 or 5 mu M CdCl2 resulted in up to 400% higher expression of genes encoding three isozymes of plant-type PEPCs and up to 200% higher expression of genes encoding two isozymes of PEPC kinase, especially in roots. The amount of PEPC and its phosphorylation status also increased by up to 40% with Cd treatment, as did enzyme activity (up to 200%). AtPPC1 and AtPPC3 transcript levels in the mutant lines were as much as 50% lower than in wild type plants, even in control (no Cd) plants. AtPPC2 expression was unaffected in both mutant lines. In general, concentrations of organic acid anions increased or remained unchanged in response to Cd treatment in each plant line, although roots of atppc1 and atppc3 mutants under control conditions had 25-50% less malate and citrate. Our results confirm the importance of root-specific (i) AtPPC3 expression for the normal development of Arabidopsis, as well as (ii) AtPPC1 and AtPPC3 upregulation for acclimation to Cd stress.