Effect of Chlorate Treatment on Nitrate Reductase and Nitrite Reductase Gene Expression in Arabidopsis thaliana

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Abstract
The herbicide chlorate has been used extensively to isolate mutants that are defective in nitrate reduction. Chlorate is a substrate for the enzyme nitrate reductase (NR), which reduces chlorate to the toxic chlorite. Because NR is a substrate (NO(3) (-))-inducible enzyme, we investigated the possibility that chlorate may also act as an inducer. Irrigation of ammonia-grown Arabidopsis plants with chlorate leads to an increase in NR mRNA in the leaves. No such increase was observed for nitrite reductase mRNA following chlorate treatment; thus, the effect seems to be specific to NR. The increase in NR mRNA did not depend on the presence of wild-type levels of NR activity or molybdenum-cofactor, as a molybdenum-cofactor mutant with low levels of NR activity displayed the same increase in NR mRNA following chlorate treatment. Even though NR mRNA levels were found to increase after chlorate treatment, no increase in NR protein was detected and the level of NR activity dropped. The lack of increase in NR protein was not due to inactivation of the cells' translational machinery, as pulse labeling experiments demonstrated that total protein synthesis was unaffected by the chlorate treatment during the time course of the experiment. Chlorate-treated plants still retain the capacity to make functional NR because NR activity could be restored by irrigating the chlorate-treated plants with nitrate. The low levels of NR protein and activity may be due to inactivation of NR by chlorite, leading to rapid degradation of the enzyme. Thus, chlorate treatment stimulates NR gene expression in Arabidopsis that is manifested only at the mRNA level and not at the protein or activity level.