title: Flux control of sulphate assimilation in Arabidopsis thaliana: adenosine 5'-phosphosulphate reductase is more susceptible than ATP sulphurylase to negative control by thiols.
abstract: The effect of externally applied L-cysteine and glutathione (GSH) on ATP sulphurylase and adenosine 5'-phosphosulphate reductase (APR), two key enzymes of assimilatory sulphate reduction, was examined in Arabidopsis thaliana root cultures. Addition of increasing L-cysteine to the nutrient solution increased internal cysteine, gamma-glutamylcysteine and GSH concentrations, and decreased APR mRNA, protein and extractable activity. An effect on APR could already be detected at 0.2 mm L-cysteine, whereas ATP sulphurylase was significantly affected only at 2 mm L-cysteine. APR mRNA, protein and activity were also decreased by GSH at 0.2 mm and higher concentrations. In the presence of L-buthionine-S, R-sulphoximine (BSO), an inhibitor of GSH synthesis, 0.2 mm L-cysteine had no effect on APR activity, indicating that GSH formed from cysteine was the regulating substance. Simultaneous addition of BSO and 0.5 mm GSH to the culture medium decreased APR mRNA, enzyme protein and activity. ATP sulphurylase activity was not affected by this treatment. Tracer experiments using (35)SO(4)(2-) in the presence of 0.5 mm L-cysteine or GSH showed that both thiols decreased sulphate uptake, APR activity and the flux of label into cysteine, GSH and protein, but had no effect on the activity of all other enzymes of assimilatory sulphate reduction and serine acetyltransferase. These results are consistent with the hypothesis that thiols regulate the flux through sulphate assimilation at the uptake and the APR step. Analysis of radioactive labelling indicates that the flux control coefficient of APR is more than 0.5 for the intracellular pathway of sulphate assimilation. This analysis also shows that the uptake of external sulphate is inhibited by GSH to a greater extent than the flux through the pathway, and that the flux control coefficient of APR for the pathway, including the transport step, is proportionately less, with a significant share of the control exerted by the transport step.
authors.affiliation: Institute of Plant Sciences, University of Berne, Altenbergrain 21, CH-3013 Berne, Switzerland.
authors.forename: Pierre
authors.initials: P
authors.lastname: Vauclare
authors.forename: Stanislav
authors.initials: S
authors.lastname: Kopriva
authors.forename: David
authors.initials: D
authors.lastname: Fell
authors.forename: Marianne
authors.initials: M
authors.lastname: Suter
authors.forename: Liliane
authors.initials: L
authors.lastname: Sticher
authors.forename: Peter
authors.initials: P
authors.lastname: von Ballmoos
authors.forename: Urs
authors.initials: U
authors.lastname: Krähenbühl
authors.forename: Roel Op
authors.initials: RO
authors.lastname: den Camp
authors.forename: Christian
authors.initials: C
authors.lastname: Brunold
mesh_terms: D017360:Arabidopsis / Q000201:enzymology* / Q000378:metabolism; D046508:Culture Techniques; D003545:Cysteine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D050862:Oxidoreductases Acting on Sulfur Group Donors; D018517:Plant Roots / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D018520:Plant Shoots / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D013430:Sulfate Adenylyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013431:Sulfates / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; D013431:Sulfates; D013438:Sulfhydryl Compounds; D010088:Oxidoreductases; D050862:Oxidoreductases Acting on Sulfur Group Donors; C019618:adenylylsulfate reductase; D013430:Sulfate Adenylyltransferase; D003545:Cysteine