Research articleMembers of a small family of nodulin-like genes are regulated under iron deficiency in roots of Arabidopsis thaliana
Highlights
► Five nodulin-like Arabidopsis genes belong to the CCC1-like gene family. ► Expression of 4 of these nodulin-like genes was positively correlated with Fe supply. ► Promoter-driven GUS expression was positively correlated with Fe supply. ► Nodulin-like T-DNA mutants showed altered Fe distribution in planta. ► A role for these nodulin-like genes in Fe storage was proposed.
Introduction
The importance of Fe as a nutritional element is explained by its crucial role in primary metabolism and stands in juxtaposition to its restricted availability in the environment [1]. The mechanisms of response to Fe deficiency and the homeostatic regulation of the Fe concentration in the cell have been frequently reviewed in the recent literature e.g.[2], [3]. In spite of the intense efforts to elucidate the molecular mechanisms that are responsible for sensing and signaling Fe deficiency, it is currently not possible to answer basic questions of how Fe deficiency is sensed by and communicated throughout plants. The response of plants to Fe deficiency involves both an increased capacity for Fe uptake [1] and a more efficient use of available Fe [4], [5]. In recent publications, we have investigated rapid changes in the Arabidopsis transcriptome in response to Fe deficiency [6], [7]. Prominent in the list of regulated transcripts are transport proteins that presumably regulate the metal concentration in cellular compartments by increasing the Fe uptake capacity into the cell or by decreasing Fe storage. As is expected, the transcripts encoding the Fe2+ transporters, AtIRT1 [8] and AtIRT2 [9], and the corresponding ferric-chelate reductases, AtFRO2 [10] and AtFRO3 [11], are increased within 6 h of Fe-deficient growth. Additionally, transcripts for an oligopeptide transporter (AtOPT3) [12] and an amino acid transporter (At5g38820) are increased in parallel with AtIRT1 and AtIRT2. Although the function of these later proteins with regard to Fe homeostasis is not known, they may catalyze transport of Fe–peptide complexes across the membrane or transport of the amino acids needed for synthesis of these peptides. Finally, transcripts for the vacuolar Fe efflux carrier, AtNRAMP4, are also increased under Fe deficiency [13], [14]. With the assumption that transcriptional changes are reflected in altered Fe fluxes, the effect of this adaptation would be to increase cytoplasmic Fe in root cells.
In contrast to transcriptional changes that indicated increased Fe uptake as an adaptation to Fe deficiency, a number of changes were observed for transporters that were not involved in Fe transport. A rapid induction was observed for MTPc3 and MTPa2 [15], [16], which catalyzed cationic metal export from the cell and Zn2+ uptake into the vacuole, respectively. The Zn2+ plasma membrane influx carrier, AtZIP3, was decreased under Fe deficiency [6], while transcript abundance increased for AtCOPT2, a plasma membrane Cu+ transporter [17]; for AtZIF1 [16], a presumptive vacuolar Zn2+-chelate transporter; and for AtIREG1 and AtIREG2 [18]. These responses to Fe deficiency could be understood in terms of regulation of an imbalance in Zn2+, Ni2+, Cu+, and/or Co2+ homeostasis in the Fe-deficient cell [18], [19]. Under Fe deficiency, an increased capacity of Zn2+ uptake, for example, might have resulted in potentially toxic concentrations of Zn in the cytoplasm. In support of this hypothesis, there was a correlation between Fe deficiency and increased shoot concentrations of Zn, Mn and Co [20].
Although much attention has been directed to transcript induction in response to Fe deficiency, we have also identified three nodulin-like genes whose transcript abundance was repressed under Fe deficiency [6], [7]. The encoded proteins were of potential importance to Fe homeostasis, because they contained a CCC1-like domain. CCC1 was initially discovered in yeast as a vacuolar Fe transporter [21], [22]. In Arabidopsis, AtVIT1 encoded a CCC1 homolog that catalyzed Fe uptake into the vacuole [23]. If the functions of these three nodulin-like proteins were similar to VIT1, then the decreased abundance of these transcripts would have been associated with a decreased capacity of the cell to store Fe in a cellular compartment.
In this report we have investigated the function of a small family of five nodulin-like proteins that all contained a CCC1-like domain and showed distant homology to AtVIT1. We presented evidence that some members of this family may participate in regulation of Fe homeostasis in plants.
Section snippets
Standardized response of nodulin-like genes to Fe deficiency
In previous reports [6], [7] we have identified a group of three so-called nodulin-like genes, whose transcript abundance was repressed under conditions of Fe-deficient growth. In an effort to identify further, related genes, the Arabidopsis genome was searched for homologies to the deduced amino acid sequences of these three nodulin-like genes. In total five genes were identified. For the purpose of this report, they were referred to as: nodulin-like1 (At1g21140), nodulin-like2 (At1g76800),
Expression of the nodulin-like gene family by Fe supply
Based on a similar response to Fe deficiency and sequence homology, we have identified a small family of nodulin-like genes whose expression was positively correlated to the Fe concentration in the growth media. Combined results from DNA microarray analyses, semi-quantitative, reverse transcription PCR and from promoter-GUS fusion constructs showed a consistent pattern of transcriptional regulation for nodulin-like1, nodulin-like2 and nodulin-like21 (Fig. 1, Fig. 2, Fig. 3, Fig. 4). In
Plant material
Arabidopsis seeds were surface-sterilized by immersion in 1.5% (v/v) NaOCl for 15 min and rinsed four times in sterile water. Seeds were vernalized for 3 d at 4 °C in the dark. Arabidopsis thaliana L. Col-0 was grown hydroponically in a growth chamber at a constant relative humidity of 75%, under a 10/14 h day/night cycle (Osram E40/ES Plantastar, 300 μmol cm−2s−1) at 21 °C (day) or 18 °C (night). Two to four seeds were sown in 1.5 ml Eppendorf® tubes that had been previously filled with 1.5 ml
Acknowledgments
We thank members of the Buckhout and Schmidt laboratories, in particular Susanne Olstowski-Jacoby and Marion Dewender, whose dedication and support were an essential part of this research effort.
References (35)
- et al.
Homing in on iron homeostasis in plants
Trends. Plant Sci.
(2009) - et al.
Time to pump iron: iron-deficiency-signaling mechanisms of higher plants
Curr. Opin. Plant Biol.
(2008) - et al.
Iron utilization and metabolism in plants
Curr. Opin. Plant Biol.
(2007) - et al.
Insights into metabolism obtained from microarray analysis
Curr. Opin. Plant Biol.
(2003) - et al.
AtFer4 ferritin is a determinant of iron homeostasis in Arabidopsis thaliana heterotrophic cells
J. Plant Physiol.
(2010) - et al.
AtIREG2 encodes a tonoplast transport protein involved in iron-dependent nickel detoxification in Arabidopsis thaliana roots
J. Biol. Chem.
(2006) - et al.
CCC1 is a transporter that mediates vacuolar iron storage in yeast
J. Biol. Chem.
(2001) - et al.
Bathophenanthrolinedisulphonic acid and bathocuproinedisulphonic acid, water soluble reagents for iron and copper
Talanta
(1961) - et al.
Changes in the proteomic and metabolic profiles of Beta vulgaris root tips in response to iron deficiency and resupply
BMC Plant Biol.
(2010) - et al.
Early iron-deficiency-induced transcriptional changes in Arabidopsis roots as revealed by microarray analyses
BMC Genomics
(2009)
Transcriptional profiling of the Arabidopsis iron deficiency response reveals conserved transition metal homeostasis networks
Plant Physiol.
A novel iron-regulated metal transporter from plants identified by functional expression in yeast
Proc. Natl. Acad. Sci. U S A
Arabidopsis IRT2 cooperates with the high-affinity iron uptake system to maintain iron homeostasis in root epidermal cells
Planta
Genetic evidence that induction of root Fe(III) chelate reductase activity is necessary for iron uptake under iron deficiency
Plant J.
The Arabidopsis AtOPT3 protein functions in metal homeostasis and movement of iron to developing seeds
Plant Physiol.
Mobilization of vacuolar iron by AtNRAMP3 and AtNRAMP4 is essential for seed germination on low iron
EMBO J.
Functional characterization of NRAMP3 and NRAMP4 from the metal hyperaccumulator Thlaspi caerulescens
New Phytol.
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