Dataset Information

Conserved Cu-MicroRNAs in Arabidopsis thaliana Function in Copper Economy under Deficiency.

ABSTRACT: Copper (Cu) is a micronutrient for plants. Three small RNAs, which are up-regulated by Cu deficiency and target transcripts for Cu proteins, are among the most conserved microRNAs in plants. It was hypothesized that these Cu-microRNAs help save Cu for the most essential Cu-proteins under deficiency. Testing this hypothesis has been a challenge due to the redundancy of the Cu microRNAs and the properties of the regulatory circuits that control Cu homeostasis. In order to investigate the role of Cu-microRNAs in Cu homeostasis during vegetative growth, we used a tandem target mimicry strategy to simultaneously inhibit the function of three conserved Cu-microRNAs in Arabidopsis thaliana. When compared to wild-type, transgenic lines that express the tandem target mimicry construct showed reduced Cu-microRNA accumulation and increased accumulation of transcripts that encode Cu proteins. As a result, these mimicry lines showed impaired photosynthesis and growth compared to wild type on low Cu, which could be ascribed to a defect in accumulation of plastocyanin, a Cu-containing photosynthetic electron carrier, which is itself not a Cu-microRNA target. These data provide experimental support for a Cu economy model where the Cu-microRNAs together function to allow maturation of essential Cu proteins under impending deficiency.

SUBMITTER: Shahbaz M

PROVIDER: S-EPMC6631055 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Conserved Cu-MicroRNAs in <i>Arabidopsis thaliana</i> Function in Copper Economy under Deficiency.

Shahbaz Muhammad M Pilon Marinus M

Plants (Basel, Switzerland) 20190529 6

Copper (Cu) is a micronutrient for plants. Three small RNAs, which are up-regulated by Cu deficiency and target transcripts for Cu proteins, are among the most conserved microRNAs in plants. It was hypothesized that these Cu-microRNAs help save Cu for the most essential Cu-proteins under deficiency. Testing this hypothesis has been a challenge due to the redundancy of the Cu microRNAs and the properties of the regulatory circuits that control Cu homeostasis. In order to investigate the role of C ...[more]

PMID: 31146453

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Project description:BackgroundThe Arabidopsis SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE (SPL) transcription factor SPL7 reprograms cellular gene expression to adapt plant growth and cellular metabolism to copper (Cu) limited culture conditions. Plant cells require Cu to maintain essential processes, such as photosynthesis, scavenging reactive oxygen species, cell wall lignification and hormone sensing. More specifically, SPL7 activity promotes a high-affinity Cu-uptake system and optimizes Cu (re-)distribution to essential Cu-proteins by means of specific miRNAs targeting mRNA transcripts for those dispensable. However, the functional mechanism underlying SPL7 activation is still to be elucidated. As SPL7 transcript levels are largely non-responsive to Cu availability, post-translational modification seems an obvious possibility. Previously, it was reported that the SPL7 SBP domain does not bind to DNA in vitro in the presence of Cu ions and that SPL7 interacts with a kin17 domain protein to raise SPL7-target gene expression upon Cu deprivation. Here we report how additional conserved SPL7 protein domains may contribute to the Cu deficiency response in Arabidopsis.ResultsCytological and biochemical approaches confirmed an operative transmembrane domain (TMD) and uncovered a dual localisation of SPL7 between the nucleus and an endomembrane system, most likely the endoplasmic reticulum (ER). This new perspective unveiled a possible link between Cu deficit and ER stress, a metabolic dysfunction found capable of inducing SPL7 targets in an SPL7-dependent manner. Moreover, in vivo protein-protein interaction assays revealed that SPL7 is able to homodimerize, probably mediated by the IRPGC domain. These observations, in combination with the constitutive activation of SPL7 targets, when ectopically expressing the N-terminal part of SPL7 including the SBP domain, shed some light on the mechanisms governing SPL7 function.ConclusionsHere, we propose a revised model of SPL7 activation and regulation. According to our results, SPL7 would be initially located to endomembranes and activated during ER stress as a result of Cu deficiency. Furthermore, we added the SPL7 dimerization in the presence of Cu ions as an additional regulatory mechanism to modulate the Cu deficiency response.

Project description:Copper and iron are essential micronutrients for most living organisms because they participate as cofactors in biological processes including respiration, photosynthesis and oxidative stress protection. In many eukaryotic organisms, including yeast and mammals, copper and iron homeostases are highly interconnected; however such interdependence is not well established in higher plants. Here we propose that COPT2, a high-affinity copper transport protein, functions under copper and iron deficiencies in Arabidopsis thaliana. COPT2 is a plasma membrane protein that functions in copper acquisition and distribution. Characterization of the COPT2 expression pattern indicates a synergic response to copper and iron limitation in roots. We have characterized a knockout of COPT2, copt2-1, that leads to increased resistance to simultaneous copper and iron deficiencies, measured as reduced leaf chlorosis and improved maintenance of the photosynthetic apparatus. We propose that COPT2 expression could play a dual role under Fe deficiency. First, COPT2 participates in the attenuation of copper deficiency responses driven by iron limitation maybe aimed to minimize further iron consume. On the other hand, global expression analyses of copt2-1 mutants versus wild type Arabidopsis plants indicate that low phosphate responses are increased in copt2-1 plants. In this sense, COPT2 function under Fe deficiency counteracts low phosphate responses. These results open up new biotechnological approaches to fight iron deficiency in crops. Four biological replicates of Arabidopsis seedlings were generated for 2 genotypes, Col-0 and copt2-1 mutant; and 3 growth condictions; first one an iron(Fe) and copper(Cu) sufficient medium (+Fe + Cu), second one an Fe deficient and Cu sufficient medium (-Fe+Cu) and third one an Fe and Cu deficient medium (-Fe-Cu). For each growth one comparasion was made, copt2-1 mutant versus Col-0; in each comparasion four biological replicates were made, two replicas were labeled with Cy5 for the mutant sample and Cy3 for the Col-0 sample, while the other two replicas were reversed-labeled.

Dataset Information

Conserved Cu-MicroRNAs in Arabidopsis thaliana Function in Copper Economy under Deficiency.

Publications

Conserved Cu-MicroRNAs in <i>Arabidopsis thaliana</i> Function in Copper Economy under Deficiency.

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