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Wilt-induced ABA biosynthesis, gene expression and down-regulation of rbcS mRNA levels in Arabidopsis thaliana
Authors:J. Williams  M. P. Bulman  S. J. Neill
Affiliation:Dept of Biological Sciences, UWE, Bristol, Coldharbour Lane, Bristol BS16 1QY, UK.
Abstract:The kinetics of wilt-induced abscisic acid (ABA) biosynthesis were investigated in shoots of Arabidopsis thaliana (L.) Heynh Landsberg erecta. ABA concentrations were measured using a radioimmunoassay (RIA) based on the monoclonal antibody MAC 252, and the RIA validated by comparison with combined gas chromatography-mass spectrometry using a [2H3] labelled internal standard. The basal ABA content of Arabidopsis shoots was ca 10 ng g?1 fresh weight; the concentrations had increased ca 4-fold within 30 min of the initiation of wilting, increased ca 8-fold after 4 h and 11-fold after 8 h. This stress-induced ABA production was dependent on de novo gene expression; pre-treatment of leaves and shoots with the metabolic inhibitors cordycepin and cycloheximide reduced the rate of subsequent stress-induced ABA biosynthesis from 12.5 ng g?1 h?1 to 1 ng g?1 h?1 and 0 ng g?1 h?1, respectively. In vitro translation of mRNA isolated from shoots subjected to wilting or ABA treatment followed by one-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoresis revealed only minor changes. The effects of wilting and ABA on the content of total ribulose 1,5-bisphosphate carboxylase/oxygenase small sub-unit (rbcS) mRNA were also determined. Both wilting and exogenous ABA resulted in a substantial reduction in the amount of rbcS mRNA, an effect readily reversed by rehydration of wilted shoots. However, the effects of wilting were not mediated solely by newly-synthesised endogenous ABA, as wilting also reduced rbcS mRNA levels in the ABA-deficient aba-1 mutant, which did not produce ABA in response to loss of turgor. The amount of rbcS mRNA was higher in aba-1 shoots, suggesting that cellular rbcS mRNA levels are normally down-regulated by ABA. Cold treatment induced ABA production in wild type shoots only, but resulted in an increased rbcS mRNA content of both wild type and aba-1 shoots.
Keywords:Abscisic acid    Arabidopsis thaliana    biosynthesis    drought    gene expression    mRNA    radioimmunoassay    ribulose 1    bisphosphate carboxylase/oxygenase
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