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Because of its ability to catalyse oxidation/reduction reactions, iron (Fe) is an essential microelement in living beings. However, high levels of Fe in the cell can lead to detrimental effects, as this metal can also catalyse the production of harmful reactive oxygen species. Therefore, tight regulation of cellular Fe concentration is required. In this thesis, two independent genes with a putative role in Fe homeostasis were characterized in the model organism Arabidopsis thaliana. AtRAI1 is homolog to the yeast Rai1p (Rat1 Interacting protein 1), an activator of 5´-to-3´ exoribonucleases, enzymes that degrade decapped RNAs. In Arabidopsis protoplasts, AtRAI1 co-localized with the decapping enzyme DCP2 (Decapping 2) in processing bodies, which are structures where mRNAs are processed for degradation. DK9, a loss-of function mutant for AtRAI1, had phenotypic traits attributable to impaired Fe homeostasis (chlorosis, reduced size) and to homeostasis of the phytohormone auxin (curly leaves and low fertility). Assessment of the activity of the Fe-dependent anti-oxidant enzymes catalase and SOD (SuperOxide Dismutase), multi-element analysis and determination of transcript levels of genes involved in Fe homeostasis showed that AtRAI1 is an upstream regulator of Fe uptake and transport in A. thaliana. The studied protein probably activates the cytosolic exoribonuclease XRN4, which is a component of the ethylene signalling pathway that is known to control Fe uptake in roots. Since homozygous knockout alleles of the analysed gene were not viable, it is concluded that AtRAI plays an essential role for Fe acquisition and proper growth of A. thaliana. The second gene studied in this thesis is BTS2 (BRUTUS2). It encodes a protein that shares similarity with BTS, an E3 ubiquitin ligase with putative roles in Fe regulation. A loss-of-function allele was more tolerant to excess Zn, a condition that leads to physiological Fe deficiency in A. thaliana. Under this condition, the mutant accumulated less Fe in the root than wild-type plants, indicating that the protein probably induces Fe uptake under Fe deficiency conditions. Furthermore, low Zn concentrations and reduced transcript levels of marker genes for excess Zn in shoots suggest an additional function in Zn translocation to the shoot. Therefore, BTS2 may play a critical role in positively regulating Fe deficiency response in roots and Zn translocation to the shoot by targeting negative regulators for degradation. |
