RNA-seq analysis of the effect of kanamycin and the ABC transporter AtWBC19 on Arabidopsis thaliana seedlings reveals changes in metal content.

Ayalew Mentewab,Morayo Adebiyi,Kinnari Matheson,Brianna Elston,Shanice Robinson,John R Battista

doi:10.1371/journal.pone.0109310

Abstract

Plants are exposed to antibiotics produced by soil microorganisms, but little is known about their responses at the transcriptional level. Likewise, few endogenous mechanisms of antibiotic resistance have been reported. The Arabidopsis thaliana ATP Binding Cassette (ABC) transporter AtWBC19 (ABCG19) is known to confer kanamycin resistance, but the exact mechanism of resistance is not well understood. Here we examined the transcriptomes of control seedlings and wbc19 mutant seedlings using RNA-seq analysis. Exposure to kanamycin indicated changes in the organization of the photosynthetic apparatus, metabolic fluxes and metal uptake. Elemental analysis showed a 60% and 80% reduction of iron uptake in control and wbc19 mutant seedlings respectively, upon exposure to kanamycin. The drop in iron content was accompanied by the upregulation of the gene encoding for FERRIC REDUCTION OXIDASE 6 (FRO6) in mutant seedlings but not by the differential expression of other transport genes known to be induced by iron deficiency. In addition, wbc19 mutants displayed a distinct expression profile in the absence of kanamycin. Most notably the expression of several zinc ion binding proteins, including ZINC TRANSPORTER 1 PRECURSOR (ZIP1) was increased, suggesting abnormal zinc uptake. Elemental analysis confirmed a 50% decrease of zinc content in wbc19 mutants. Thus, the antibiotic resistance gene WBC19 appears to also have a role in zinc uptake.

Highlights

Plants are exposed to a variety of antibiotics produced by soil microorganisms
RNA-seq analysis was conducted to capture the effect of kanamycin on Arabidopsis seedlings and examine how the response was modulated by WBC19
To compensate for the confounding effect that might be added by the neomycin phosphotransferase type II (NPTII) gene, a line with an insertion of the same T-DNA, but where no gene is disrupted was used as control

Summary

Introduction

Plants are exposed to a variety of antibiotics produced by soil microorganisms. One such antibiotic is kanamycin, originally isolated from the soil bacterium Streptomyces kanamyceticus. Kanamycin belongs to the aminoglycoside class of antibiotics. Aminoglycosides directly bind to the A site of the 16S rRNA of prokaryotic ribosomes causing codon misreading and inhibiting translocation [1,2,3]. The presence of an adenosine at position 1408 of the 16S rRNA is a key feature of prokaryotic ribosomes that allows binding of aminoglycosides. Based on the similarity of their ribosomes to those of prokaryotes, plastids and mitochondria may represent the primary targets of aminoglycoside antibiotics in plants

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Conclusion