Sucrose induction of Arabidopsis miR398 represses two Cu/Zn superoxide dismutases
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MicroRNAs (miRNAs) are ∼21-nt RNAs that reduce target accumulation through mRNA cleavage or translational repression. Arabidopsis miR398 regulates mRNAs encoding two copper superoxide dismutase (CSD) enzymes and a cytochrome c oxidase subunit. miR398 itself is down-regulated in response to copper and stress. Here we show that miR398 is positively regulated by sucrose, resulting in decreased CSD1 and CSD2 mRNA and protein accumulation. This sucrose regulation is maintained both in the presence and absence of physiologically relevant levels of supplemental copper. Additionally, we show that plants expressing CSD1 and CSD2 mRNAs with altered miR398 complementarity sites display increased mRNA accumulation, whereas CSD1 and CSD2 protein accumulation remain sensitive to miR398 levels, suggesting that miR398 can act as a translational repressor when target site complementarity is reduced. These results reveal a novel miR398 regulatory mechanism and demonstrate that plant miRNA targets can resist miRNA regulation at the mRNA level while maintaining sensitivity at the level of protein accumulation. Our results suggest that even in plants, where miRNAs are thought to act primarily through target mRNA cleavage, monitoring target protein levels along with target mRNA levels is necessary to fully assess the consequences of disrupted miRNA–mRNA pairing. Moreover, the limited complementarity required to maintain robust miR398-directed repression of target protein accumulation suggests that similarly regulated endogenous plant miRNA targets may have eluded detection.
KeywordsmiR398 microRNA Post-transcriptional regulation Sucrose regulation Superoxide dismutase
Copper superoxide dismutase
Iron superoxide dismutase
Plant nutrient medium
Plant nutrient medium without micronutrients
Reactive oxygen species
We thank Daniel Kliebenstein for providing the anti-CSD1, anti-CSD2, and anti-FSD antibodies, the Salk Institute Genomic Analysis Laboratory for generating the sequence-indexed T-DNA insertion mutants, and the Arabidopsis Biological Resource Center at Ohio State University for seeds, cDNA clones, and BAC clones. We thank Matthew Lingard, Lucia Strader, and Andrew Woodward for critical comments on the manuscript, and David Bartel for helpful discussions. This research was supported by the National Institutes of Health (R24-GM069512), the G. Harold and Leila Y. Mathers Charitable Foundation, and the Robert A. Welch Foundation (C-1309). D.V.D. was supported in part by a training grant from the National Institutes of Health (T32-GM08362).
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